Photoinduced inverse Sonogashira coupling reaction

Alkynes are widely used in chemistry, medicine and materials science. Here we demonstrate a transition-metal and photocatalyst-free inverse Sonogashira coupling reaction between iodoalkynes and (hetero)arenes or alkenes under visible-light irradiation. Mechanistic and computational studies suggest that iodoalkynes can be directly activated by visible light irradiation, and an excited state iodoalkyne acted as an “alkynyl radical synthetic equivalent”, reacting with a series of C(sp²)–H bonds for coupling products. This work should open new windows in radical chemistry and alkynylation method.

A transition-metal and photocatalyst-free, photoinduced inverse Sonogashira coupling reaction was developed. Under visible-light irradiation, the excited state iodoalkyne acted as an “alkynyl radical synthetic equivalent”.

Alkynes are among the most important class of compounds in organic chemistry. Because of their structural rigidity, special electronic properties and numerous methods available for the functionalization of the triple bond, alkynes are important tools and structural elements both in medicinal chemistry and materials sciences.¹ Therefore, the development of a new methodology to introduce carbon–carbon triple bonds is of great importance in organic chemistry. The Sonogashira coupling reaction is typically used for the formation of C(sp)–C(sp²) bonds starting from hetero(aryl) halides and terminal alkynes.² Recently, “inverse Sonogashira coupling” involving the direct alkynylation of unreactive C(sp²)–H bonds with readily available alkynyl halides has received growing interest in the development of a complementary strategy (Fig. 1a). Various main-group and transition metals have been developed to promote this transformation.³ In addition, a photomediated Sonogashira reaction without a photocatalyst was also developed by several groups (Fig. 1b).⁴Open in a separate windowFig. 1Models of alkynylation. (a) Conventional inverse Sonogashira reaction. (b) Photomediated Sonogashira reaction. (c) SOMOphilic alkynylation. (d) Photoinduced inverse Sonogashira reaction.In recent years, SOMOphilic alkylnylation (SOMO = singly occupied molecular orbital) has become an excellent method of introducing alkynyl groups (Fig. 1c).⁵ Based on photoredox and transition metal catalysis, numerous in situ generated radicals undergo α-addition and β-elimination to alkynyl reagents, like the broadly applicable ethynylbenziodoxolone (EBX) reagent. Various radical alkynylations were thus discovered by Li,⁶ Chen,⁷ Waser,⁸ and many other groups.⁹ However, extending the scope of radical precursors, more atom–economic reactions, and a deeper understanding of the mechanism in these transformations are still highly desirable.After the discovering of trityl radicals by Gomberg in 1900, the “rational” era of radical chemistry has since begun.¹⁰ Now, the development of radical reactions, especially those involving C(sp³) and C(sp²) radicals, enables rapid access to drug discovery, agrochemistry, materials science, and other disciplines.¹¹ However, the C(sp) radical remains a baffling species. Due to their very high energy, short life time, and limited and harsh preparation methods, alkynyl radicals remain an elusive species, which just exists in some extreme environments, like outer-space and the petrochemical industry.¹² Even though alkynyl radicals have been proposed as intermediates for some alkynylation methods, they were regarded as mysterious species and ignored by organic chemists for a long time.¹³ Recently, two approaches have been developed to aid the alkynyl radical generation step. In 2015, Hashmi and collaborators reported a [Au₂(μ-dppm)₂]²⁺ catalyzed free radical–radical C(sp)–C(sp³) bond coupling reaction between iodoalkynes and aliphatic amines.¹⁴ Under irradiation of sunlight, the dimeric gold complex was proposed to reduce the iodine acetylide to an alkynyl radical. In 2017, Li developed a transition-metal-free alkynylation reaction between iodoalkyne and 2-indolinone.¹⁵ Iodoalkynes could release alkynyl radicals under high temperature conditions. In 2019, we reported an Au(i) and Ir(iii) catalyzed alkynylative cyclization of o-alkylnylphenols with iodoalkynes, wherein the photosensitized energy transfer promoted the oxidative addition of a gold(i) complex with iodoalkynes.¹⁶ Based on our continuous interest in haloalkyne and photo-chemistry, we proposed that an iodoalkyne could be a potential “alkynyl radical precursor” under light irradiation. In this work, we uncovered a novel mode of transition-metal and photocatalyst-free, direct photoexcitation of iodoalkynes for the inverse Sonogashira coupling reaction with arenes, heteroarenes, and alkenes via an “alkynyl-radical type” transfer (Fig. 1d).     

Controlled monodefluorination and alkylation of C(sp3)–F bonds by lanthanide photocatalysts: importance of metal–ligand cooperativity

The controlled functionalization of a single fluorine in a CF₃ group is difficult and rare. Photochemical C–F bond functionalization of the sp³-C–H bond in trifluorotoluene, PhCF₃, is achieved using catalysts made from earth-abundant lanthanides, (Cp^Me4)₂Ln(2-O-3,5-^tBu₂-C₆H₂)(1-C{N(CH)₂N(ⁱPr)}) (Ln = La, Ce, Nd and Sm, Cp^Me4 = C₅Me₄H). The Ce complex is the most effective at mediating hydrodefluorination and defluoroalkylative coupling of PhCF₃ with alkenes; addition of magnesium dialkyls enables catalytic C–F bond cleavage and C–C bond formation by all the complexes. Mechanistic experiments confirm the essential role of the Lewis acidic metal and support an inner-sphere mechanism of C–F activation. Computational studies agree that coordination of the C–F substrate is essential for C–F bond cleavage. The unexpected catalytic activity for all members is made possible by the light-absorbing ability of the redox non-innocent ligands. The results described herein underscore the importance of metal–ligand cooperativity, specifically the synergy between the metal and ligand in both light absorption and redox reactivity, in organometallic photocatalysis.

The controlled functionalization of a single fluorine in a CF₃ group is difficult and rare. Photochemical C–F bond functionalization is achieved using catalysts made from a range of earth-abundant lanthanides by using a ligand that enables M–L cooperativity.

Photoredox catalysis is a powerful synthetic method for the functionalization of inert molecules using single electron transfer (SET) reactivity^1–3 under irradiation with visible light.⁴ This has enabled challenging transformations under mild conditions including C–H activation,^5–7 radical cross-coupling,^8–11 and the valorization of lignin.^12,13 However, detailed mechanistic studies of photoredox systems are difficult due to their inherent complexity and the short lifetimes of photoexcited intermediates.Many lanthanides are more abundant in the environment than copper and their salts are less toxic than those of iron, so their potential for applications in catalysis merits exploration.^14–17 In 1990, divalent Sm, Eu, and Yb complexes Ln(Cp*)₂ (Cp* = C₅Me₅), were shown to more efficiently cleave vinylic C–F bonds when photolyzed, stoichiometrically forming Ln(iii) halide complexes, and suggesting the value of increasing the reducing power of the Ln^II excited state.¹⁸ Subsequently, analogous reactions to cleave the weaker C–Cl and C–Br bonds could be made catalytic in Ln(ii) halide (Ln = Sm, Eu, Yb), under near UV-photolysis conditions, by the addition of sacrificial reductant such as Zn or Al.^19,20 The addition of simple donor ligands enabled benzylic C–Cl cleavage by Eu^II under blue light irradiation.²¹ The addition of an organic photocatalyst or a photo-absorbing substrate to Lewis acidic LnX₃ salts (X = halide, triflate) has also been used to enhance the catalysis.²² Ln centers (Ln = Nd, Dy, Lu) with light-absorbing ligands such as porphyrins or phthalocyanins have been used to stoichiometrically dechlorinate phenols.²³Few reports of lanthanide photoredox catalysis exist with Ce^III complexes receiving the most attention. Ce possesses both an accessible III/IV redox couple and an allowed excitation from the 4f¹ ground state to the 5d¹ excited state, which can give rise to luminescent behaviour. It is also the cheapest and most readily isolated of the rare earths, offering a promising alternative to current precious metal photocatalysts.Building on the pioneering work on stoichiometric photoluminescent Ce chemistry,^24,25 in 2015 Schelter and co-workers demonstrated the utility of Ce^III in photocatalysis.^26,27 Their Ce^III amido complexes were catalysts for chlorine atom abstraction from benzyl chloride (Fig. 1, top), with both NaN(SiMe₃)₂ and additional Ce⁰ required for turnover.²⁸Open in a separate windowFig. 1Previous examples of photocatalytic C–X (X = halide) bond cleavage, and this work.They proposed an inner-sphere mechanism involving Ce⋯ClCR₃ adduct formation that provides an additional thermodynamic driving force to a bond cleavage that was otherwise out of range of the reducing power of the Ce excited state. A more sterically congested Ce^III tris(guanidinate) operates via an outer-sphere single electron transfer (SET) mechanism to cleave aryl iodides,²⁸ highlighting the mechanistic diversity that is possible in these systems.²⁹To date, ligands that support lanthanide-centered photocatalysts have been limited to halides, pseudohalides, and simple N-donors.^30,31 No organometallic lanthanide photocatalyst has yet been reported that combines the photoexcitable Ce cation with multidentate, tunable ligands. We have developed organometallic lanthanide complexes as sustainable catalysts,^16,17,32,33 and considered that those capable of forming an inner-sphere adduct, and absorbing light, could achieve the unusual and difficult, selective catalytic conversion of strong sp³ C–F bonds.Fluorine forms the strongest single bond to carbon and the C–F bond is ca. 25 kcal mol⁻¹ stronger than the C–Cl bond in monohaloalkanes, and the C–H bond in alkanes.³⁴ The selective activation and functionalization of C–F bonds is important, both due to the high bioaccumulation and toxicity of many perfluorinated compounds,³⁵ and the utility of fluorinated pharmaceuticals.³⁶ However, stoichiometric C(sp³)–F bond activation reactions are rare.^37–40 In particular, it is difficult to facilitate the controlled cleavage of a single C–F bond as the C(sp³)–F bond strength decreases as each F is removed and the remaining C–F bonds lengthen.^41,42This obstacle makes a radical methodology more attractive.^43–49 Jui and co-workers have demonstrated that some common photocatalysts can selectively activate a single C–F bond to form the putative ArCF₂˙ radical, which can either be quenched directly via H atom transfer (HAT), or coupled with an alkene followed by HAT to generate difluoroalkanes (Fig. 1, middle).^50,51 Gschwind and König have shown the photochemical functionalization of electron-poor trifluoromethylarenes.⁵² Nishimoto and Yasuda have described related C–F coupling protocols of perfluoroalkylarenes using tin reagents and an iridium photocatalyst.⁵³Here we show how selective, catalytic C–F bond functionalization can be achieved using a new family of Ln^III compounds supported by a light-absorbing aryloxide-tethered N-heterocyclic carbene, Cp^Me4, and pseudohalide ligands (Fig. 1, lower). We show that visible light-irradiated Ce complexes can selectively abstract a single fluoride from PhCF₃ and catalyze its alkylation by MgR₂ to afford PhCF₂R. The PhCF₂˙ can also be quenched to selectively form PhCF₂H or further alkylated via coupling with an alkene or other metal alkyls. We use combined experiment and density functional theory (DFT) computations to show the importance of coordination of the fluorinated substrate to the Lewis acidic metal in C–F activation, and the utility of the ligand in enabling photoredox catalysis for other lanthanide congeners.     

Correction: Influence of the primary and secondary coordination spheres on nitric oxide adsorption and reactivity in cobalt(ii)–triazolate frameworks

Correction for ‘Influence of the primary and secondary coordination spheres on nitric oxide adsorption and reactivity in cobalt(ii)–triazolate frameworks’ by Julia Oktawiec et al., Chem. Sci., 2021, DOI: 10.1039/d1sc03994f.

The authors regret that incorrect details were given for ref. 35, 37 and 59 in the original article. The correct versions of ref. 35, 37 and 59 are given below as ref. 1, 2 and 3, respectively.The Royal Society of Chemistry apologises for these errors and any consequent inconvenience to authors and readers.     

Computational design of an amidase by combining the best electrostatic features of two promiscuous hydrolases

While there has been emerging interest in designing new enzymes to solve practical challenges, computer-based options to redesign catalytically active proteins are rather limited. Here, a rational QM/MM molecular dynamics strategy based on combining the best electrostatic properties of enzymes with activity in a common reaction is presented. The computational protocol has been applied to the re-design of the protein scaffold of an existing promiscuous esterase from Bacillus subtilis Bs2 to enhance its secondary amidase activity. After the alignment of Bs2 with a non-homologous amidase Candida antarctica lipase B (CALB) within rotation quaternions, a relevant spatial aspartate residue of the latter was transferred to the former as a means to favor the electrostatics of transition state formation, where a clear separation of charges takes place. Deep computational insights, however, revealed a significant conformational change caused by the amino acid replacement, provoking a shift in the pK_a of the inserted aspartate and counteracting the anticipated catalytic effect. This prediction was experimentally confirmed with a 1.3-fold increase in activity. The good agreement between theoretical and experimental results, as well as the linear correlation between the electrostatic properties and the activation energy barriers, suggest that the presented computational-based investigation can transform in an enzyme engineering approach.

A computational strategy, based on combining the best electrostatic properties of enzymes with activity on a common reaction, is presented and applied to the re-design of the protein scaffold of an promiscuous esterase to enhance its secondary amidase activity.

The application of enzymes for desired chemical transformations has been demonstrated by the report of novel and functional designed structures.^1–5 Recent advances in molecular biology and screening technologies have enabled the creation of enzymes via directed evolution. By mimicking the process of natural evolution, iterative cycles of (semi-)random mutations facilitate the improvement of proteins in the laboratory through screening and selection, and hence the identification of active variants.^6–12 Minimal structural information is needed for this strategy and distal sites critical for enzyme catalysis can also be identified. Nevertheless, directed evolution is limited by the fact that, even with the most efficient high-throughput system, only a fraction of all the possible mutants of a given enzyme can be sampled within a set timeframe.¹³ Furthermore, the development of an efficient screening system for a tailored reaction remains challenging. Recently machine-learning (ML) methods have been proposed to expedite evolution and expand the number of properties that can be optimized.^14,15 However, in order to create enzymes with novel reactivities by means of ML methods, protein engineers will have to use proteins with sequences not assigned to the designated reaction or with properties other than those of specific interest, which currently is a technical challenge. Sequence–function data from engineering experiments must be collected to catalogue the natural diversity of proteins in order to convert ML into a useful tool.¹⁵An alternative approach is a rational design, a technique that modifies selected residues at specific positions of an already existing protein scaffold through the analysis of existing mechanistic and structural data.¹⁶ To reveal the structures of the protein in the full catalytic process under physiological conditions, including metastable transition state (TS) structures, computer simulations are essential. Among all the computer-assisted design strategies, two philosophies can be identified: the redesign of the active site of an existing substrate-promiscuous enzyme and the de novo design that constructs an enzyme “from scratch”. The use of promiscuous enzymes is found to be a very promising starting point for the design of new and highly efficient biocatalysts.^17,18 However, the knowledge about the particular molecular mechanisms that allow enzymes to catalyze more than one chemical reaction is still under debate.^19–22Because both the enzyme redesign and the de novo design approach require knowledge of the TS of the reaction to be catalyzed, quantum mechanical (QM) calculations offer crucial complementary information that accelerates the development of novel designed reactions. Moreover, multiscale methods are the only tool that can offer a detailed atomistic picture of the reactions in the active site of the enzyme, which can be dramatically different from that in the gas phase or solution. In multiscale methods, electrons of the reacting fragments are explicitly described by QM methods and the large and complex interacting environment (the fully solvated protein) is described by molecular mechanics (MM) force fields. The mechanism of a reaction in the active site of an enzyme can be determined within these hybrid QM/MM methods through the extensive exploration of the Free Energy Surface (FES). This allows the determination of the rate-limiting step in a multi-step process and, within the framework of Transition State Theory (TST),²³ the prediction of rate constants directly comparable with experiments. Previous studies combining computer simulations with experimental kinetic measurements have demonstrated the good agreement that can be achieved,^24,25 which obviously depends on the quality of both simulations and experiments. In this regard, the error in the determination of activation free energies associated with the use of computational methods such as the umbrella sampling method,^26,27 employed in the present study, is usually accepted to be within 1 kcal mol⁻¹.²⁸Optimizing the secondary activity of promiscuous enzymes is a non-trivial challenge as can be illustrated by analysing the Bacillus subtilis esterase Bs2. While Bs2 is recognized as a serine hydrolase whose primary reaction is the hydrolysis of esters, it can also catalyze the hydrolysis of the amide bond of N-(4-nitrophenyl)-butyramide as a secondary reaction (Fig. 1).^22,29 Previously directed evolution experiments by Arnold and co-workers resulted in a 7-mutation variant with a 100-fold enhancement of the esterase activity (using para-nitrobenzyl butyrate as the substrate).³⁰ Bornscheuer and co-workers used a combination of directed evolution and rational design based on docking and classical energy minimization to get a 3-fold increase of the amidase activity of Bs2 after two single mutations.²² In a larger context, despite the successes of different computer-assisted designs of new enzymes, it has been argued that the high activities of the best artificial enzymes have been largely due to directed evolution and the contribution of computation was comparatively modest.³¹Open in a separate windowFig. 1Schematic representation of the reaction mechanism of the hydrolysis of N-(4-nitrophenyl)-butyramide catalyzed by Bs2. (a) Acylation step: the nucleophilic addition of Ser189 to the carbonyl followed by the breaking of the C–N bond is triggered by His399-assisted proton shuffling, and the leaving group, in this case, is 4-nitroaniline. (b) Hydrolysis step: the nucleophilic addition of a water molecule followed by the resolution of the acyl–enzyme complex is triggered by His399-assisted protein shuffling, yielding butyric acid as a product.We envisaged that creating mutations to optimize the preorganization of the protein environment will result in a variant that exhibits improved activity for the desired reaction.³² Based on our recent QM/MM studies of different enzymatic reactions, we have quantified and shown how the reactivity of different proteins can be rationalized from their electrostatic properties,^{24,25,33–36} as the pioneering studies reported by Warshel and co-workers.^37–39 The computed changes of the electrostatic potential or the electric field exerted by the studied proteins on the key atoms of the substrates reflect that there is a small reorganization of these entities when evolving from the reactant state (RS) to the TS at the lowest energy cost.^24,33–36 The electrostatic effects within the active site of the enzyme, therefore, appear to be critical for the electronic reorganization of the reactants during chemical transformations. These studies support the idea that the electrostatic properties of enzymes are the origin of their catalytic features;⁴⁰ consequently, we view that a detailed understanding of the molecular mechanism, including the evolution of electrostatic potential generated in the active site of the enzyme, could be useful in future computer-assisted protein design methods.To engineer enzymes with optimal electrostatic preorganization, comparative analysis between unrelated natural enzymes that catalyze the same chemical reactions can be a reliable strategy. In previous studies, we have shown that Candida antarctica lipase B (CALB) also displays amidase activity similar to that of Bs2, though being non-homologous with each other.^24,25 QM/MM studies of the amidase reaction catalyzed by wild-type Bs2 and CALB enzymes were previously conducted.^24,25 We expect that the favorable features of each enzyme could be isolated and combined to create a redesigned enzyme with improved catalytic activity for the secondary amidase reaction. In the present paper, based on our knowledge derived from previous comparative studies, and by applying the concept of electrostatic pre-organization,^{24,33–36,40–43} a variant with improved activity for the designated amidase reaction was generated. After overlapping the structures of both proteins in one of the located TSs, through the use of a rotation quaternion around selected atoms of the substrate, a catalytically improved Bs2 variant was delineated. In particular, residues of Bs2 with an unfavorable electrostatic effect on catalysis were substituted by those placed in an equivalent spatial position in CALB with a favorable effect, as explained in detail below. The QM/MM FES of the full catalytic reaction in the proposed variant, combined with the experimental characterization, will be used to propose a general computer-based strategy that can be potentially used to design new enzymes.     

Correction: The oxygen-resistant [FeFe]-hydrogenase CbA5H harbors an unknown radical signal

Correction for ‘The oxygen-resistant [FeFe]-hydrogenase CbA5H harbors an unknown radical signal’ by Melanie Heghmanns et al., Chem. Sci., 2022, 13, 7289–7294, https://doi.org/10.1039/D2SC00385F.

The authors realized that incorrect references were cited following the sentence “In conjunction with the signal''s significant width, the frequency dependence clearly indicates spin–spin interaction between the F-clusters.” The correct references are shown below as ref. 1 and 2.Additionally ref. 36 and 37 were reversed in the reference list. The correct ref. 36 is shown below as ref. 3 and the correct ref. 37 is shown below as ref. 4.The Royal Society of Chemistry apologises for these errors and any consequent inconvenience to authors and readers.     

Stereoselective tandem iridium-catalyzed alkene isomerization-cope rearrangement of ω-diene epoxides: efficient access to acyclic 1,6-dicarbonyl compounds

The Cope rearrangement of 2,3-divinyloxiranes, a rare example of epoxide C–C bond cleavage, results in 4,5-dihydrooxepines which are amenable to hydrolysis, furnishing 1,6-dicarbonyl compounds containing two contiguous stereocenters at the 3- and 4-positions. We employ an Ir-based alkene isomerization catalyst to form the reactive 2,3-divinyloxirane in situ with complete regio- and stereocontrol, which translates into excellent control over the stereochemistry of the resulting oxepines and ultimately to an attractive strategy towards 1,6-dicarbonyl compounds.

Iridium catalyzed alkene isomerization-cope rearrangement of ω-diene epoxide furnishes 3,4-dihydrooxepines. These oxepines are hydrolyzed to diastereomerically pure 1,6-dicarbonyl compound containing two contiguous stereocenters within acyclic system.

1,6-Dicarbonyl compounds are widespread as targets and intermediates in organic synthesis.¹ Due to the “dissonant” polarizing effect induced by the two carbonyl groups,² these motifs are challenging to retrosynthetically disconnect into classical synthons. Unsurprisingly, many approaches toward 1,6-dicarbonyls rely on dimerization of α,β-unsaturated carbonyl compounds (Scheme 1a)³ or oxidative cleavage of substituted cyclohexene derivatives⁴ which significantly limits the range of possible products. Alternative strategies, such as the ring-opening of donor–acceptor cyclopropanes with enolate nucleophiles, efficiently form the 1,6-dicarbonyl skeleton, albeit with limited substrate scope (Scheme 1b).⁵ The Cope rearrangement of 1,5-dienes, featuring oxygen functionality in the 3- and 4-positions,⁶ represents a promising strategy towards 1,6-dicarbonyl compounds but suffers from lack of stereocontrol over the diene substrates, resulting in diastereomeric mixtures of products (Scheme 1c).Open in a separate windowScheme 1Selected approaches towards the formation of 1,6-dicarbonyl compounds and our proposed approach.A conceptually related approach towards the preparation of 1,6-dicarbonyl compounds is through the hydrolysis of 3,4-dihydrooxepines (Scheme 1d), which are in turn generated through the Cope rearrangement of 2,3-divinyloxiranes.⁷ Such a sigmatropic rearrangement is also noteworthy as a rare example where an epoxide C–C bond is selectively cleaved over the usually more reactive C–O bond. This intriguing rearrangement has been studied but its use in synthesis is scarce, presumably due to difficulties in the stereoselective synthesis and handling of the key divinyl epoxides.In line with our interest in the strategic application of alkene isomerization to generate reactive synthetic intermediates in stereodefined form,⁸ we posited to form the reactive 2,3-divinyloxiranes in situ, through alkene isomerization^9,10 of the simpler allyl epoxides, which are accessible in enantiomerically enriched form.¹¹ Such a strategy might greatly facilitate access to these intermediates and therefore uncover a synthetically attractive route toward 1,6-dicarbonyl compounds featuring two contiguous stereocenters.With this idea in mind, we first explored the isomerization and subsequent Cope rearrangement of allyl-vinyl epoxides 1 (Scheme 2). To induce isomerization, we employed a cationic iridium-based catalytic system,¹² which is known to reliably isomerize alkenes with high degrees of regio- and stereocontrol.¹³Open in a separate windowScheme 2Substrate scope for the tandem iridium-catalyzed alkene isomerization-Cope rearrangement of allyl-vinyl epoxides.In line with our expectations, our model substrate 1a (R² = R³ = H, R⁴ = Me, R⁵ = CO₂Et) was smoothly isomerized at 65 °C in the presence of 1.5 mol% of Ir dimer to obtain the corresponding divinyl epoxide with a complete E-selectivity. With suitable conditions for alkene isomerization in hand, we exposed substrate 1a to the Ir-based catalytic system at 120 °C and were equally pleased to observe the 4,5-dihydrooxepine product 2a, resulting from the tandem isomerization-Cope rearrangement as a single diastereoisomer in 81% yield. We proceeded to test the generality of our protocol with respect to different alkene and epoxide substitution patterns. Pleasingly, product 2b was generated with complete stereoselectivity, showcasing the compatibility of the reaction conditions with potentially labile tertiary stereocenters α to the ester group. We then wondered whether the anti-diastereomer could be accessed starting from the corresponding cis allyl-vinyl epoxide. Indeed, in line with the known stereospecific behavior of the Cope rearrangement, we obtained the complementary diastereomer 2c. Turning our attention to more highly substituted epoxides, we were pleased to observe the formation of dihydrooxepines 2d and 2e, which correspond to 1,6-keto-aldehyde and diketone products, respectively. Substrate 1f (R² = R⁴ = R⁵ = H, R³ = Ph), which features an unactivated vinyl group, also underwent the rearrangement, demonstrating that an activated alkenyl group is not required for a successful outcome. Similarly, product 2g featuring two alkyl groups is also generated, with high diastereoselectivity albeit in moderate yield. Products featuring ethyl and methyl ester 2h, 2i could also be obtained in good yields and diastereoselectivity. We next tested substrate 1j (R² = Me, R³ = Ph, R⁴ = CH₂CH₂Ph, R⁵ = H), as a geometric-mixture of the double bond (E : Z = 1.1 : 1) and in accordance with the stereospecificity of the process, the oxepine 2j was obtained as a mixture of two diastereomers with the same ratio. Disappointingly, substrate 1k did not undergo isomerization, presumably due to the Lewis basic nature of the ketone, likely poisoning the Ir-catalyst.During our study, we noticed that allyl-vinyl epoxides bearing electron donating groups on the vinyl moiety tend to decompose during purification by column chromatography on silica gel. This obstacle further motivated us to explore diallyl epoxides 3 as substrates, where the reactive divinyl epoxide would be generated by isomerization of both allyl fragments. Notably, these diallyl epoxides are much more stable compared to their vinyl counterparts and can be readily prepared in two steps from simple alkynes.¹⁴ To our delight, diallyl epoxide 3a (R = CH₂OMe) smoothly underwent the double isomerization-Cope rearrangement cascade at 140 °C, furnishing oxepine 2l with impressive yield and diastereoselectivity (Scheme 3). The use of alkene isomerization to form the reactive divinyl epoxide in situ avoids the isolation of the unstable divinyl epoxide, while controlling the stereochemistry of both double bonds, particularly not trivial to achieve using classical olefination reactions. Products 2m and 2n feature ester and silyl groups, highlighting the functional group tolerance of the catalytic system.Open in a separate windowScheme 3Substrate scope for tandem iridium-catalyzed double alkene isomerization-Cope rearrangement of diallyl epoxides.Our next objective was to hydrolyze the diastereomerically pure oxepines obtained through the rearrangement in a stereoretentive fashion, revealing the acyclic 1,6-dicarbonyl motif. Pleasingly, diversely substituted oxepines 2 underwent smooth hydrolysis either using 5 mol% of Pd(MeCN)₂Cl₂¹⁵ at 50 °C or an acidic aqueous solution to form 1,6-dicarbonyls 4 in diastereomerically pure form (Scheme 4).¹⁶ Dicarbonyl products featuring labile tertiary centers 4a and 4b are formed under these conditions with excellent diastereoselectivities and yields. Without surprise, oxepine 2f (R² = R⁴ = R⁵ = H, R³ = Ph) furnished the keto-substituted product 4c in good yield. The relative stereochemistry of 4b was unambiguously confirmed by single crystal X-ray diffraction analysis of the corresponding carboxylic acid 7 (Scheme 4b).¹⁷ The reaction is scalable to ½ gram of substrate and could be performed in a single-pot operation without isolation of the intermediate oxepine (Scheme 4b). By using this approach, 1h provides 4b in 61% yield as a single diastereomer, underlining the synthetic potential and efficiency of this method.Open in a separate windowScheme 4Hydrolysis of oxepines and one-pot sequence.     

Electrooxidative o-carborane chalcogenations without directing groups: cage activation by copper catalysis at room temperature

Copper-catalyzed electrochemical direct chalcogenations of o-carboranes was established at room temperature. Thereby, a series of cage C-sulfenylated and C-selenylated o-carboranes anchored with valuable functional groups was accessed with high levels of position- and chemo-selectivity control. The cupraelectrocatalysis provided efficient means to activate otherwise inert cage C–H bonds for the late-stage diversification of o-carboranes.

Copper-catalyzed electrochemical cage C–H chalcogenation of o-carboranes has been realized to enable the synthesis of various cage C-sulfenylated and C-selenylated o-carboranes.

Carboranes are polyhedral molecular boron–carbon clusters, which display unique properties, such as a boron enriched content, icosahedron geometry and three-dimensional electronic delocalization.¹ These features render carboranes as valuable building blocks for applications to optoelectronics,² as nanomaterials, in supramolecular design,³ organometallic coordination chemistry,⁴ and boron neutron capture therapy (BNCT) agents.⁵ As a consequence, considerable progress has been witnessed in transition metal-catalyzed regioselective cage B–H functionalization of o-carboranes⁶ and different functional motifs have been incorporated into the cage boron vertices.^7–10 However, progress in this research arena continues to be considerably limited by the shortage of robust and efficient methods to access carborane-functionalized molecules. While C–S bonds are important structural motifs in various biologically active molecules and functional materials,¹¹ strategies for the assembly of chalcogen-substituted carboranes continue to be scarce. A major challenge is hence represented by the strong coordination abilities of thiols to most transition metals, which often lead to catalyst deactivation.¹² While copper-catalyzed B(4,5)–H disulfenylation of o-carboranes was achieved,^7e elevated reaction temperature was required, and 8-aminoquinoline was necessary as bidentate directing group. The bidentate directing group¹³ needs to be installed and removed, which jeopardizes the overall efficacy. Likewise, an organometallic strategy was recently devised for cysteine borylation with a stoichiometric platinum(ii)-based carboranes.¹⁴ Meanwhile, oxidative cage B/C–H functionalizations largely call for noble transition metal catalysts¹⁵ and stoichiometric amounts of chemical oxidants, such as expensive silver(i) salts.¹⁶In recent years, electricity has been identified as an increasingly viable, sustainable redox equivalent for environmentally-benign molecular synthesis.^17,18 While significant advances have been realized by the merger of electrocatalysis with organometallic bond activation,¹⁹ electrochemical carborane functionalizations continue unfortunately to be underdevelopment. In sharp contrast, we have now devised a strategy for unprecedented copper-catalyzed electrochemical cage C–H chalcogenations of o-carboranes in a dehydrogenative manner, assembling a variety of C-sulfenylated and C-selenylated o-carboranes (Fig. 1a). It is noteworthy that our electrochemical cage C–S/Se modification approach is devoid of chemical oxidants, and does not need any directing groups, operative at room temperature.Open in a separate windowFig. 1Electrochemical diversification of o-carboranes and optimization of reaction conditions. ^aReaction conditions: procedure A: 1a (0.10 mmol), 2a (0.3 mmol), CuOAc (15 mol%), 2-PhPy (15 mol%), LiOtBu (0.2 mmol), TBAI (2.0 equiv.), solvent (3 mL), platinum cathode (10 mm × 15 mm × 0.25 mm), graphite felt (GF) anode (10 mm × 15 mm × 6 mm), 2 mA, under air, r.t., 16 h. ^bYield was determined by ¹H NMR with CH₂Br₂ as the internal standard. ^cIsolated yields in parenthesis. ^dKI (1.0 equiv.) as additive. ^eProcedure B: 2 (0.3 mmol), LiOtBu (0.2 mmol), TBAI (2.0 equiv.), solvent (3.0 mL), 2 mA, r.t., 3 h, then adding 1a (0.10 mmol), 2-PhPy (15 mol%), CuOAc (15 mol%), 2 mA, rt, 16 h. ^f2b (0.3 mmol), LiOtBu (0.2 mmol), KI (1.0 equiv.), TBAI (2.0 equiv.), solvent (3.0 mL), 2 mA, r.t., 3 h, then adding 1a (0.10 mmol), 2-PhPy (15 mol%), CuOAc (15 mol%), r.t., 16 h. TBAI = tetrabutylammonium iodide, TBAPF₆ = tetrabutylammonium hexafluorophosphate. DCE = 1,2-dichloroethane, THF = tetrahydrofuran.We commenced our studies by probing various reaction conditions for the envisioned copper-catalyzed cage C–H thiolation of o-carborane in an operationally simple undivided cell setup equipped with a GF (graphite felt) anode and a Pt cathode (Fig. 1b and Table S1^†). After extensive experimentation, we observed that the thiolation of substrate 1 proceeded efficiently with catalytic amounts of CuOAc and 2-phenylpyridine, albeit in the presence of 2 equivalents LiOtBu as the base, and 2 equivalents n-Bu₄NI as the electrolyte at room temperature under a constant current of 2 mA (entry 1). The yield was reduced when other copper sources or additives were used (entries 2–5). Surprisingly, n-Bu₄NPF₆ as the electrolyte failed to facilitate the carborane modification, indicating that n-Bu₄NI operates not only as electrolyte, but also as a redox mediator (entry 6). Altering the stoichiometry of the electrolyte or using KI did not improve the performance (entries 7–8). Product formation was not observed, when the reaction was conducted with DCE as the solvent, while CH₃CN resulted in a drop of the catalytic performance (entries 9–10). Control experiments confirmed the essential role of the electricity and the catalyst (entries 11–12), while a sequential procedure was found to be beneficial (entries 13–15).With the optimized reaction conditions in hand, we explored the versatility of the cage C–H thiolation of o-carborane 1a with different thiols 2 (Scheme 1). Electron-rich as well as electron-deficient substituents on the arenes were found to be amenable to the electrocatalyzed C–H activation, providing the corresponding thiolation products 3aa–3ao in good to excellent yields. Thereby, a variety of synthetically useful functional groups, such as fluoro (3ae, 3am), chloro (3af, 3ak, 3an) and bromo (3ag, 3al), were fully tolerated, which should prove instrumental for further late-stage manipulations. Various disubstituted aromatic and heterocyclic thiols afforded the corresponding cage C–S modified products 3ap–3as. Notably, aliphatic thiols efficiently underwent the electrochemical transformation to provide the corresponding cage alkylthiolated products 3at–3au. Notably, the halogen-containing thiols (2e–2f, 2k–2n and 2q) reacted selectively with o-carboranes to deliver the desired products without halide coupling byproducts being observed. The connectivity of the products 3aa, 3am and 3ao was unambiguously verified by X-ray single crystal diffraction analysis.²²Open in a separate windowScheme 1Electrochemical C–H thiolation of o-carborane 1a. (a) Procedure B. (b) KI (1 equiv.). (c) Cul as the catalyst.Encouraged by the efficiency of the cupraelectro-oxidative cage C–H thiolation, we became intrigued to explore the chalcogenantion of differently-decorated o-carboranes 1 (Scheme 2). Electronically diverse carboranes 1 served as competent coupling partners, giving the corresponding thiolation products 4bo–4do with high levels of efficacy in position-selective manner. The strategy was not restricted to phenyl-substituted o-carboranes. Indeed, substrates bearing benzyl and even alkyl groups also performed well to deliver the desired products 4eo–4ga. It is noteworthy that the C–H activation approach was also compatible with selenols to give the o-carboranes 4av–4fv. The molecular structures of the carborane 4br and 4av were unambiguously verified by single-crystal X-ray diffraction.²²Open in a separate windowScheme 2Electrochemical cage C–H chalcogenation of o-carboranes. (a) Procedure B. (b) KI (1 equiv.).Scaffold functionalization of the thus obtained carborane 3ag provided the alkynylated derivative 5a and amine 5b (Scheme 3), giving access to carborane-based host materials of relevant to phosphorescent organic light-emitting diodes.²⁰Open in a separate windowScheme 3Late-stage diversification.Next, we became attracted to delineating the mode of the cupraelectro-catalyzed cage C–H chalcogenation. To this end, control experiments were performed (Scheme 4a). First, electrocatalysis in the presence of TEMPO or Ph₂C CH₂ gave the desired product 3aa. EPR studies of thiol 2a, LiOtBu and THF under the electrochemical conditions showed a small radical signal, which might be attributed to a thiol radical.²¹ Second, the cupraelectrocatalysis occurred efficiently in the dark. Third, detailed cyclovoltammetric analysis of the thiol and iodide mediator (Scheme 4b and ESI^†)²¹ revealed an irreversible oxidation of the thiol anion at E_p = −0.62 V vs. Ag/Ag⁺ and two oxidation events for the iodide, including an irreversible oxidation at E_p = 0.12 V vs. Ag/Ag⁺ and a reversible oxidation at E_p = 0.44 V vs. Ag/Ag⁺, which is in good agreement with the literature reported iodide oxidation potentials,^18c,d and is suggestive of the preferential oxidation of the iodide as a redox mediator. In this context, the use of n-Bu₄NI as a redox mediator to achieve copper-catalyzed electrochemical arene C–H aminations had been documented.^18d Furthermore, we calculated the redox potential of complex C by means of DFT calculations at the PW6B95-D4/def2-TZVP + SMD(MeCN)//TPSS-D3BJ/def2-SVP level of theory.²¹ These studies revealed a calculated oxidation half-wave potential for complex C is E^o,calc_1/2 = −0.08 V vs. SCE. Hence, iodide is a competent redox mediator to achieve the transformation from complex C to complex D. Analysis of non-covalent interactions²¹ in complex C (Fig. 2) show the presence of a weak stabilization interaction between the chalcogen''s anisole group and the 2-phenylpyridine. In contrast, in complex D these interactions were found more relevant between the o-carborane phenyl group and the chalcogen aromatic motif.Open in a separate windowFig. 2Non-covalent interaction plots for the complexes C and D. Strong attractive interactions are shown in blue, weak attractive interactions are given in green, while red corresponds to repulsive interactions. Ar = 4-MeOC₆H₄.Open in a separate windowScheme 4Control experiments and cyclic voltammograms.On the basis of the aforementioned findings,¹⁸ a plausible reaction mechanism is proposed in Scheme 5, which commences with an anodic single electron-transfer (SET) oxidation of the thiol anion E to form the sulfur-centered radical F. Subsequently, the copper(i) species A reacts with the sulfur radical F to deliver copper(ii) complex B, which next reacts with o-carborane 1 in the presence of LiOtBu to generate a copper(ii)-o-carborane complex C. Thereafter, the complex C is oxidized by the anodically generated redox mediator I₂ to furnish the copper(iii) species D,^18d which subsequently undergoes reductive elimination, affording the final product and regenerating the catalytically active complex A. Alternatively, the direct oxidation of copper(ii) complex C by electricity to generate copper(iii) species D can not be excluded at this stage.^18a,bOpen in a separate windowScheme 5Proposed reaction mechanism.In conclusion, a sustainable electrocatalytic C–H chalcogenation of o-carboranes with thiols and selenols was realized at room temperature by earth abundant copper catalysis. The C–H activation was characterized by mild reaction conditions and high functional group tolerance, leading to the facile assembly of various o-carboranes. Thereby, a transformative platform for the design of cage C–S and C–Se o-carboranes was established that avoids chemical oxidants by environmentally-sound electricity in the absence of directing groups. A plausible mechanism of paired electrolysis was established by detailed mechanistic studies.     

A Paal–Knorr agent for chemoproteomic profiling of targets of isoketals in cells

Natural systems produce various γ-dicarbonyl-bearing compounds that can covalently modify lysine in protein targets via the classic Paal–Knorr reaction. Among them is a unique class of lipid-derived electrophiles – isoketals that exhibit high chemical reactivity and critical biological functions. However, their target selectivity and profiles in complex proteomes remain unknown. Here we report a Paal–Knorr agent, 4-oxonon-8-ynal (herein termed ONAyne), for surveying the reactivity and selectivity of the γ-dicarbonyl warhead in biological systems. Using an unbiased open-search strategy, we demonstrated the lysine specificity of ONAyne on a proteome-wide scale and characterized six probe-derived modifications, including the initial pyrrole adduct and its oxidative products (i.e., lactam and hydroxylactam adducts), an enlactam adduct from dehydration of hydroxylactam, and two chemotypes formed in the presence of endogenous formaldehyde (i.e., fulvene and aldehyde adducts). Furthermore, combined with quantitative chemoproteomics in a competitive format, ONAyne permitted global, in situ, and site-specific profiling of targeted lysine residues of two specific isomers of isoketals, levuglandin (LG) D2 and E2. The functional analyses reveal that LG-derived adduction drives inhibition of malate dehydrogenase MDH2 and exhibits a crosstalk with two epigenetic marks on histone H2B in macrophages. Our approach should be broadly useful for target profiling of bioactive γ-dicarbonyls in diverse biological contexts.

Natural systems produce various γ-dicarbonyl-bearing compounds that can covalently modify lysine in protein targets via the classic Paal–Knorr reaction.

Synthetic chemistry methods have been increasingly underscored by their potential to be repurposed as biocompatible methods for both chemical biology and drug discovery. The most-known examples of such a repurposing approach include the Staudinger ligation¹ and the Huisgen-based click chemistry.² Moreover, bioconjugation of cysteine and lysine can be built upon facile chemical processes,³ while chemoselective labelling of other polar residues (e.g., histidine,⁴ methionine,⁵ tyrosine,⁶ aspartic and glutamic acids^7,8) requires more elaborate chemistry, thereby offering a powerful means to study the structure and function of proteins, even at a proteome-wide scale.The classical Paal–Knorr reaction has been reported for a single-step pyrrole synthesis in 1884.^9,10 The reaction involves the condensation of γ-dicarbonyl with a primary amine under mild conditions (e.g., room temperature, mild acid) to give pyrrole through the intermediary hemiaminals followed by rapid dehydration of highly unstable pyrrolidine adducts (Fig. S1^†).Interestingly, we and others have recently demonstrated that the Paal–Knorr reaction can also readily take place in native biological systems.^11–13 More importantly, the Paal–Knorr precursor γ-dicarbonyl resides on many endogenous metabolites and bioactive natural products.¹⁴ Among them of particular interest are isoketals¹⁵ (IsoKs, also known as γ-ketoaldehydes) which are a unique class of lipid derived electrophiles (LDEs) formed from lipid peroxidation (Fig. S2^†)¹⁶ that has emerged as an important mechanism for cells to regulate redox signalling and inflammatory responses,¹⁷ and drive ferroptosis,¹⁸ and this field has exponentially grown over the past few years. It has been well documented that the γ-dicarbonyl group of IsoKs can rapidly and predominantly react with lysine via the Paal–Knorr reaction to form a pyrrole adduct in vitro (Fig. 1).¹⁵ Further, the pyrrole formed by IsoKs can be easily oxidized to yield lactam and hydroxylactam products in the presence of molecular oxygen (Fig. 1). These rapid reactions are essentially irreversible. Hence, IsoKs react with protein approximately two orders of magnitude faster than the most-studied LDE 4-hydoxynonenal (4-HNE) that contains α,β-unsaturated carbonyl to generally adduct protein cysteines by Michael addition (Fig. S3^†).¹⁵ Due to this unique adduction chemistry and rapid reactivity, IsoKs exhibit intriguing biological activities, including inhibition of the nucleosome complex formation,¹⁹ high-density lipoprotein function,²⁰ mitochondrial respiration and calcium homeostasis,²¹ as well as activation of hepatic stellate cells.²² Furthermore, increases in IsoK-protein adducts have been identified in many major diseases,²³ such as atherosclerosis, Alzheimer''s disease, hypertension and so on.Open in a separate windowFig. 1The Paal–Knorr precursor γ-dicarbonyl reacts with the lysine residue on proteins to form diverse chemotypes via two pathways. The red arrow shows the oxidation pathway, while the blue one shows the formaldehyde pathway.Despite the chemical uniqueness, biological significance, and pathophysiological relevance of IsoKs, their residue selectivity and target profiles in complex proteomes remain unknown, hampering the studies of their mechanisms of action (MoAs). Pioneered by the Cravatt group, the competitive ABPP (activity-based protein profiling) has been the method of choice to analyse the molecular interactions between electrophiles (e.g., LDEs,²⁴ oncometabolites,²⁵ natural products,^26,27 covalent ligands and drugs^28–30) and nucleophilic amino acids across complex proteomes. In this regard, many residue-specific chemistry methods and probes have been developed for such studies. For example, several lysine-specific probes based on the activated ester warheads (e.g., sulfotetrafluorophenyl, STP;³¹N-hydroxysuccinimide, NHS³²) have recently been developed to analyse electrophile–lysine interactions at a proteome-wide scale in human tumour cells, which provides rich resources of ligandable sites for covalent probes and potential therapeutics. Although these approaches can also be presumably leveraged to globally and site-specifically profile lysine-specific targets IsoKs, the reaction kinetics and target preference of activated ester-based probes likely differ from those of γ-dicarbonyls, possibly resulting in misinterpretation of ABPP competition results. Ideally, a lysine profiling probe used for a competitive ABPP analysis of IsoKs should therefore possess the same, or at least a similar, warhead moiety. Furthermore, due to the lack of reactive carbonyl groups on IsoK-derived protein adducts, several recently developed carbonyl-directed ligation probes for studying LDE-adductions are also not suitable for target profiling of IsoKs.^33,34Towards this end, we sought to design a “clickable” γ-dicarbonyl probe for profiling lysine residues and, in combination with the competitive ABPP strategy, for analysing IsoK adductions in native proteomes. Considering that the diversity of various regio- and stereo- IsoK isomers¹⁵ (a total of 64, Fig. S2^†) in chemical reactivity and bioactivities is likely attributed to the substitution of γ-dicarbonyls at positions 2 and 3, the “clickable” alkyne handle needs to be rationally implemented onto the 4-methyl group in order to minimize the biases when competing with IsoKs in target engagement. Interestingly, we reasoned that 4-oxonon-8-ynal, a previously reported Paal–Knorr agent used as an intermediate for synthesizing fatty acid probes³⁵ or oxa-tricyclic compounds,³⁶ could be repurposed for the γ-dicarbonyl-directed ABPP application. With this chemical in hand (herein termed ONAyne, Fig. 2A), we first used western blotting to detect its utility in labelling proteins, allowing visualization of a dose-dependent labelling of the proteome in situ (Fig. S4^†). Next, we set up to incorporate this probe into a well-established chemoproteomic workflow for site-specific lysine profiling in situ (Fig. 2A). Specifically, intact cells were labelled with ONAyne in situ (200 μM, 2 h, 37 °C, a condition showing little cytotoxicity, Fig. S5^†), and the probe-labelled proteome was harvested and processed into tryptic peptides. The resulting probe-labelled peptides were conjugated with both light and heavy azido-UV-cleavable-biotin reagents (1 : 1) via Cu^I-catalyzed azide–alkyne cycloaddition reaction (CuAAC, also known as click chemistry). The biotinylated peptides were enriched with streptavidin beads and photoreleased for LC-MS/MS-based proteomics. The ONAyne-labelled peptides covalently conjugated with light and heavy tags would yield an isotopic signature. We considered only those modified peptide assignments whose MS1 data reflected a light/heavy ratio close to 1.0, thereby increasing the accuracy of these peptide identifications. Using this criterium, we applied a targeted database search to profile three expected probe-derived modifications (PDMs), including 13 pyrrole peptide adducts (Δ273.15), 77 lactam peptide adducts (Δ289.14), and 557 hydroxylactam peptide adducts (Δ305.14), comprising 585 lysine residues on 299 proteins (Fig. S6 and S7^†). Among them, the hydroxylactam adducts were present predominately, since the pyrrole formed by this probe, the same as IsoKs, can be easily oxidized when being exposed to O₂. This finding was in accordance with a previous report where the pyrrole adducts formed by the reaction between IsoK and free lysine could not be detected, but rather their oxidized forms.³⁷ Regardless, all three types of adducts were found in one lysine site of EF1A1 (K387, Fig. S8^†), further confirming the intrinsic relationship among those adductions in situ.Open in a separate windowFig. 2Adduct profile and proteome-wide selectivity of the γ-dicarbonyl probe ONAyne. (A) Chemical structure of ONAyne and schematic workflow for identifying ONAyne-adducted sites across the proteome. (B) Bar chart showing the distribution of six types of ONAyne-derived modifications formed in situ and in vitro (note: before probe labelling, small molecules in cell lysates were filtered out through desalting columns).State-of-the-art blind search can offer an opportunity to explore unexpected chemotypes (i.e., modifications) derived from a chemical probe and to unbiasedly assess its proteome-wide residue selectivity.^38,39 We therefore sought to use one of such tools termed pChem³⁸ to re-analyse the MS data (see Methods, ESI^†). Surprisingly, the pChem search identified three new and abundant PDMs (Fig. 1 and Table S1^†), which dramatically expand the ONAyne-profiled lysinome (2305 sites versus 585 sites). Overall, these newly identified PDMs accounted for 74.6% of all identifications (Fig. 2B and Table S2^†). Among them, the PDM of Δ287.13 (Fig. 1 and S7^†) might be an enlactam product via dehydration of the probe-derived hydroxylactam adduct. The other two might be explained by the plausible mechanism as follows (Fig. 1). The endogenous formaldehyde (FA, produced in substantial quantities in biological systems) reacts with the probe-derived pyrrole adduct via nucleophilic addition to form a carbinol intermediate, followed by rapid dehydration to a fulvene (Δ285.15, Fig. S7^†) and immediate oxidation to an aldehyde (Δ301.14, Fig. S7^†). In line with this mechanism, the amount of FA-derived PDMs was largely eliminated when the in vitro ONAyne labelling was performed in the FA-less cell lysates (Fig. 2B and Table S3^†). Undoubtedly, the detailed mechanisms underlying the formation of these unexpected PDMs require further investigation, and so does the reaction kinetics. Regardless, all main PDMs from ONAyne predominantly target the lysine residue with an average localization probability of 0.77, demonstrating their proteome-wide selectivity (Fig. S9^†).Next, we adapted an ABPP approach to globally and site-specifically quantify the reactivity of lysine towards the γ-dicarbonyl warhead through a dose-dependent labelling strategy (Fig. 3A) that has been proved to be successful for other lysine-specific probes (e.g., STP alkyne).³¹ Specifically, MDA-MB-231 cell lysates were treated with low versus high concentrations of ONAyne (1 mM versus 0.1 mM) for 1 h. Probe-labelled proteomes were digested into tryptic peptides that were then conjugated to isotopically labelled biotin tags via CuAAC for enrichment, identification and quantification. In principle, hyperreactive lysine would saturate labelling at the low probe concentration, whereas less reactive ones would show concentration-dependent increases in labelling. For fair comparison, the STP alkyne-based lysine profiling data were generated by using the same chemoproteomic workflow. Although 77.5% (3207) ONAyne-adducted lysine sites can also be profiled by STP alkyne-based analysis, the former indeed has its distinct target-profile with 930 lysine sites newly identified (Fig. S10 and Table S4^†). Interestingly, sequence motif analysis with pLogo⁴⁰ revealed a significant difference in consensus motifs between ONAyne- and STP alkyne-targeting lysines (Fig. S11^†).Open in a separate windowFig. 3ONAyne-based quantitative reactivity profiling of proteomic lysines. (A) Schematic workflow for quantitative profiling of ONAyne–lysine reactions using the dose-dependent ABPP strategy (B) Box plots showing the distribution of R_10:1 values quantified in ONAyne- and STP alkyne-based ABPP analyses, respectively. Red lines showing the median values. ***p ≤ 0.001 two-tailed Student''s t-test. (C) Representative extracted ion chromatograms (XICs) showing changes in the EF1A1 peptide bearing K273 that is adducted as indicated, with the profiles for light and heavy-labelled peptides in blue and red, respectively.Moreover, we quantified the ratio (R_{1 mM:0.1 mM}) for a total of 2439 ONAyne-tagged lysines (on 922 proteins) and 17904 STP alkyne-tagged lysines (on 4447 proteins) across three biological replicates (Fig. S12 and Table S5^†). Strikingly, only 26.7% (651) of quantified sites exhibited nearly dose-dependent increases (R_{1 mM:0.1 mM} > 5.0) in reactivity with ONAyne, an indicative of dose saturation (Fig. 3B and C). In contrast, such dose-dependent labelling events accounted for >69.1% of all quantified lysine sites in the STP alkyne-based ABPP analysis.³¹ This finding is in accordance with the extremely fast kinetics of reaction between lysine and γ-dicarbonyls (prone to saturation). Nonetheless, by applying 10-fold lower probe concentrations, overall 1628 (80.2%) detected lysines could be labelled in a fully concentration-dependent manner with the median R_10:1 value of 8.1 (Fig. 3B, C, S12 and Table S5^†). Next, we asked whether the dose-depending quantitation data (100 μM versus 10 μM) can be harnessed to predict functionality. By retrieving the functional information for all quantified lysines from the UniProt Knowledgebase, we found that those hyper-reactive lysines could not be significantly over-represented with annotation (Fig. S12^†). Nonetheless, among all quantified lysines, 509 (25.1%) possess functional annotations, while merely 2.5% of the human lysinome can be annotated. Moreover, 381 (74.8%) ONAyne-labelled sites are known targets of various enzymatic post-translational modifications (PTMs), such as acetylation, succinylation, methylation and so on (Fig. S13^†). In contrast, all known PTM sites accounted for only 59.6% of the annotated human lysinome. These findings therefore highlight the intrinsic reactivity of ONAyne towards the ‘hot spots’ of endogenous lysine PTMs.The aforementioned results validate ONAyne as a fit-for-purpose lysine-specific chemoproteomic probe for competitive isoTOP-ABPP application of γ-dicarbonyl target profiling. Inspired by this, we next applied ONAyne-based chemoproteomics in an in situ competitive format (Fig. 4A) to globally profile lysine sites targeted by a mixture of levuglandin (LG) D₂ and E₂, two specific isomers of IsoKs that can be synthesized conveniently from prostaglandin H₂ (ref. 41) (Fig. S2^†). Specifically, mouse macrophage RAW264.7 cells (a well-established model cell line to study LDE-induced inflammatory effects) were treated with 2 μM LGs or vehicle (DMSO) for 2 h, followed by ONAyne labelling for an additional 2 h. The probe-labelled proteomes were processed as mentioned above. For each lysine detected in this analysis, we calculated a control/treatment ratio (R_C/T). Adduction of a lysine site by LGs would reduce its accessibility to the ONAyne probe, and thus a higher R_C/T indicates increased adduction. In total, we quantified 2000 lysine sites on 834 proteins across five biological replicates. Among them, 102 (5.1%) sites exhibited decreases of reactivity towards LGs treatment (P < 0.05, Table S6^†), thereby being considered as potential targets of LGs. Notably, we found that different lysines on the same proteins showed varying sensitivity towards LGs (e.g., LGs targeted K3 of thioredoxin but not K8, K85 and K94, Table S6^†), an indicative of changes in reactivity, though we could not formally exclude the effects of changes in protein expression on the quantified competition ratios. Regardless, to the best of our knowledge, the proteome-wide identification of potential protein targets by IsoKs/LGs has not been possible until this work.Open in a separate windowFig. 4ONAyne-based in situ competitive ABPP uncovers functional targets of LGs in macrophages. (A) Schematic workflow for profiling LGs–lysine interactions using ONAyne-based in situ competitive ABPP. (B) Volcano plot showing the log₂ values of the ratio between the control (heavy) and LGs-treated (light) channels and the −log₁₀(P) of the statistical significance in a two-sample t-test for all quantified lysines. Potential targets of LGs are shown in blue (R_C/T>1.2, P < 0.05), with the validated ones in red. (C) Bar chart showing the inhibitory effect of 2 μM LGs on the cellular enzymatic activity of MDH2. Data represent means ± standard deviation (n = 3). Statistical significance was calculated with two-tailed Student''s t-tests. (D) Pretreatment of LGs dose-dependently blocked ONAyne-labelling of MDH2 in RAW264.7 cells, as measured by western blotting-based ABPP. (E and F) LGs dose-dependently decreased the H2BK5 acetylation level in RAW 264.7 cells, as measured either by western blotting (E) or by immunofluorescence imaging (F). n = 3. For G, nuclei were visualized using DAPI (blue).We initially evaluated MDH2 (malate dehydrogenase, mitochondrial, also known as MDHM), an important metabolic enzyme that possesses four previously uncharacterized liganded lysine sites (K157, K239, K301 and K329, Fig. 4B) that are far from the active site (Fig. S14^†). We found that LGs dramatically reduced the catalytic activity of MDH2 in RAW264.7 cells (Fig. 4C), suggesting a potentially allosteric effect. We next turned our attention to the targeted sites residing on histone proteins, which happen to be modified by functionally important acetylation, including H2BK5ac (Fig. 4B) that can regulate both stemness and epithelial–mesenchymal transition of trophoblast stem cells.⁴² We therefore hypothesized that rapid adduction by LGs competes with the enzymatic formation of this epigenetic mark. Immunoblotting-based competitive ABPP confirmed that LGs dose-dependently blocked probe labelling of H2B (Fig. 4D). Further, both western blots and immunofluorescence assays revealed that LG treatment decreased the level of acetylation of H2BK5 (average R_C/T = 1.3, P = 0.007) in a concentration-dependent manner (Fig. 4E and F). Likewise, a similar competitive crosstalk was observed between acetylation and LG-adduction on H2BK20 (average R_C/T = 1.2, P = 0.01) that is required for chromatin assembly⁴³ and/or gene regulation⁴⁴ (Fig. 4B and S15^†). Notably, these findings, together with several previous reports by us and others about histone lysine ketoamide adduction by another important LDE, 4-oxo-2-noenal,^11,45,46 highlight again the potentially important link between lipid peroxidation and epigenetic regulation. In addition to the targets validated as above, many other leads also merit functional studies considering diverse biological or physiologic effects of LGs in macrophages.     

One-electron bonds in copper–aluminum and copper–gallium complexes

Odd-electron bonds have unique electronic structures and are often encountered as transiently stable, homonuclear species. In this study, a pair of copper complexes supported by Group 13 metalloligands, M[N((o-C₆H₄)NCH₂PⁱPr₂)₃] (M = Al or Ga), featuring two-center/one-electron (2c/1e) σ-bonds were synthesized by one-electron reduction of the corresponding Cu(i) ⇢ M(III) counterparts. The copper bimetallic complexes were investigated by X-ray diffraction, cyclic voltammetry, electron paramagnetic spectroscopy, and density functional theory calculations. The combined experimental and theoretical data corroborate that the unpaired spin is delocalized across Cu, M, and ancillary atoms, and the singly occupied molecular orbital (SOMO) corresponds to a σ-(Cu–M) bond involving the Cu 4p_z and M ns/np_z atomic orbitals. Collectively, the data suggest the covalent nature of these interactions, which represent the first examples of odd-electron σ-bonds for the heavier Group 13 elements Al and Ga.

Hanging on by a thread. Formally zerovalent copper complexes with an Al(iii) or Ga(iii) support were investigated. The combined experimental and theoretical data corroborate the presence of an odd-electron σ-bond between Cu and the Group 13 center.

Odd-electron σ-bonds, where the electrons are delocalized between two atoms, can occur as two-center/one-electron (2c/1e) or two-center/three-electron (2c/3e) interactions. Proposed by Pauling in 1931,¹ odd-electron σ-bonds have garnered attention because of their fundamental importance to chemical bonding and their relationship to radical species generated during oxidative stress in biological systems.^2–14 Examples of compounds exhibiting odd-electron bonding are typically homonuclear (like H₂⁺, He₂⁺, and alkali metal dimers) and transiently stable, limiting them to spectroscopic characterization.^1,11,15–18The first solid-state structure of a formally one-electron σ-bond was a tetraphosphabenzene species (Fig. 1a) which was formed by the coupling of two diphosphirenyl radicals.¹⁹ Following this discovery, the formation of discrete 2c/1e σ-bonds, where the odd-electron is delocalized between two homonuclear main group centers, was reported for B·B and then extended to P·P.^8,17,20 Of note, the first solid-state structure of a B·B compound was reported in only 2014 (Fig. 1b).²¹ Examples of 2c/1e σ-bonds between the heavier Group 13 congeners are even more lacking because of the greater propensity for their unpaired spins to couple, forming larger more stable clusters.⁸ To our knowledge, there are only three structurally characterized examples of odd-electron bonds for the heavy Group 13 atoms,²² and these examples are all homonuclear π-radicals (Fig. 1c).^23–26Open in a separate windowFig. 1Select examples of structurally characterized molecules (a–d) featuring odd-electron bonds.Heteronuclear odd-electron σ-bonds are also rare. The Cu(TPB) complex, where TPB is a trisphosphinoborane, is the single structural example of a 2c/1e bond between heteroatoms (Fig. 1d).²⁷ The authors described the bonding as Cu·B, where the unpaired electron is heavily polarized toward B. A theoretical study predicted that such a bond would also exist between Cu and Al, but no heavier analogues of Cu(TPB) have been synthesized to date.²⁸ Furthermore, the heavier Group 13 elements by virtue of their lower electronegativity compared to B should facilitate greater covalent interactions with the Cu center.Hence, we sought to target formally zerovalent Cu complexes supported by Al(III) or Ga(III) as an extension of the previously reported isoelectronic nickelate species and Cu(TPB).²⁹ Herein, we describe the synthesis, structure, spectroscopic characterization, and DFT calculations of cationic [CuML]⁺ complexes (L = [N((o-C₆H₄)NCH₂PⁱPr₂)₃]³⁻; M = Al and Ga) as well as their one-electron reduced metalloradical counterparts that feature discrete 2c/1e bonds.     

Chelation-assisted C–C bond activation of biphenylene by gold(i) halides

A chelation-assisted oxidative addition of gold(i) into the C–C bond of biphenylene is reported here. The presence of a coordinating group (pyridine, phosphine) in the biphenylene unit enabled the use of readily available gold(i) halide precursors providing a new, straightforward entry towards cyclometalated (N^C^C)- and (P^C)-gold(iii) complexes. Our study, combining spectroscopic and crystallographic data with DFT calculations, showcases the importance of neighboring, weakly coordinating groups towards the successful activation of strained C–C bonds by gold.

Pyridine and phosphine directing groups promote the C–C activation of biphenylene by readily available gold(i) halides rendering a new entry to (N^C^C)- and (P^C)-gold(iii) species.

Activation of C–C bonds by transition metals is challenging given their inertness and ubiquitous presence alongside competing C–H bonds.¹ Both the intrinsic steric hindrance as well as the highly directional character of the p orbitals involved in the σ_C–C bond impose a high kinetic barrier for this type of processes.^2,3 Biphenylene, a stable antiaromatic system featuring two benzene rings connected via a four-membered cycle, has found widespread application in the study of C–C bond activation. Since the seminal report from Eisch et al. on the oxidative addition of a nickel(0) complex into the C–C bond of biphenylene,⁴ several other late transition metals have been successfully applied in this context.⁵ Interestingly, despite the general reluctance of gold(i) to undergo oxidative addition,⁶ its oxidative insertion into the C–C bond of biphenylene was demonstrated in two consecutive reports by the groups of Toste^7a and Bourissou,^7b respectively. The high energy barrier associated with the oxidation of gold could be overcome by the utilization of gold(i) precursors bearing ligands that exhibit either a strongly electron-donating character (e.g. IPr = [1,3-bis(2,6-diisopropylphenyl)imidazole-2-ylidene])^7a or small bite angles (e.g. DPCb = diphosphino-carborane).^7b,8 In line with these two approaches, more sophisticated bidentate (N^C)- and (P^N)-ligated gold(i) complexes have also been shown to aid the activation of biphenylene at ambient temperature (Scheme 1a).^7c,dOpen in a separate windowScheme 1(a) Previous reports on oxidative addition of ligated gold(i) precursors onto biphenylene. (b) This work: pyridine- and phosphine-directed C–C bond activation of biphenylene by commercially available gold(i) halides.In this context, we hypothesized that the oxidative insertion of gold(i) into the C–C bond of biphenylene could be facilitated by the presence of a neighboring chelating group.⁹ This approach would not only circumvent the need for gold(i) precursors featuring strong σ-donor or highly tailored bidentate ligands but also offer a de novo entry towards interesting, less explored ligand templates. However, recent work by Breher and co-workers showcased the difficulty of achieving such a transformation.¹⁰Herein, we report the oxidative insertion of readily available gold(i) halide precursors into the C–C bond of biphenylene. The appendage of both pyridine and phosphine donors in close proximity to the σ_C–C bond bridging the two aromatic rings provides additional stabilization to the metal center and results in a de novo entry to cyclometalated (N^C^C)- and (P^C)gold(iii) complexes (Scheme 1b).Our study commenced with the preparation of 5-chloro-1-pyridino-biphenylene system 2via Pd-catalyzed Suzuki cross coupling reaction between 2-bromo-3-methylpyridine and 2-(5-chlorobiphenylen-1-yl)-4,4,5,5-tetramethyl-1,3,2-dioxaborolane 1 (Scheme 2).¹¹ To our delight, the reaction of 2 with gold(i) iodide in toluene at 130 °C furnished complex κ³-(N^C^C)Au(iii)–I 3 in 60% yield.^12,13 Complex 3 was isolated as yellow plate-type crystals from the reaction mixture and its molecular structure was unambiguously assigned by NMR spectroscopy, high-resolution mass spectrometry (HR-MS) and crystallographic analysis. Complex 3 exhibits the expected square-planar geometry around the metal center, with a Au–I bond length of 2.6558(3) Å.¹⁴ The choice of a neutral weakly bound gold(i)-iodide precursor is key for a successful reaction outcome: similar reactions in the presence of [(NHC)AuCl + AgSbF₆] failed to deliver the desired biscyclometalation adducts, as reported by Breher et al. in ref. 10. The oxidative insertion of gold(i) iodide into the four-membered ring of pyridino-substituted biphenylene provides a novel and synthetically efficient entry to κ³-(N^C^C)gold(iii) halides. These species have recently found widespread application as precursors for the characterization of highly labile, catalytically relevant gold(iii) intermediates,^15a–d as well as for the preparation of highly efficient emitters in OLEDs.^15e–g Previous synthetic routes towards these attractive biscyclometalated gold(iii) systems involved microwave-assisted double C–H functionalization reactions that typically proceed with low to moderate yields.^15aOpen in a separate windowScheme 2Synthesis of complex 3via oxidative addition of Au(i) into the C–C bond of pyridine-substituted biphenylene. X-ray structures of complex 3 with atoms drawn using 50% probability ellipsoids. Hydrogen atoms have been omitted for clarity. Additional selected bond distances [Å]: N–Au = 2.126(2), C1–Au = 1.973(2), C2–Au = 2.025(2), Au–I = 2.6558(3) and bond angles [deg]: N–Au–I = 99.25(6), N–Au–C1 = 79.82(9), C1–Au–C2 = 81.2(1), C2–Au–I = 99.73(8). For experimental details, see ESI.^†Encouraged by the successful results obtained with the pyridine-substituted biphenylene and considering the prominent use of phosphines in gold chemistry,^6,16 we wondered whether the same reactivity would be observed for a P-containing system. To this end, both adamantyl- and tert-butyl-substituted phosphines were appended in C1 position of the biphenylene motif. Starting from 5-chlorobiphenylene-1-carbaldehyde 4, phosphine-substituted biphenylenes 5a and 5b could be accessed in 3 steps (aldehyde reduction to the corresponding alcohol, Appel reaction and nucleophilic displacement of the corresponding benzylic halide) in 64 and 57% overall yields, respectively.¹³ The reactions of 5a and 5b with commercially available gold(i) halides (Me₂SAuCl and AuI) furnished the corresponding mononuclear complexes 7a–b and 8a–b, respectively (Scheme 3).¹³ All these complexes were fully characterized and the structures of 7a, 7b and 8a were unambiguously characterized by X-ray diffraction analysis.¹³ Interestingly, the nature of the halide has a clear effect on the chemical shift of the phosphine ligand so that a Δδ of ca. 5 ppm can be observed in the ³¹P NMR spectra of 7a–b (Au–Cl) compared to 8a–b (Au–I), the latter being the more deshielded. The Au–X bond length is also impacted, with a longer Au–I distance (2.5608(1) Å for 8a) compared to that measured in the Au–Cl analogue (2.2941(7) Å for 7a) (Δd = 0.27 Å).¹³Open in a separate windowScheme 3Synthesis and reactivity of complexes 7a–b, 8a–b, 9 and 10. X-ray structure of complexes 11b, 12 and 14 with atoms drawn using 50% probability ellipsoids. Hydrogen atoms have been omitted for clarity. For experimental details and X-ray structures see ESI.^†Despite numerous attempts to promote the C–C activation in these complexes,^10,13 all reactions resulted in the formation of highly stable cationic species 11a–b and 12, which could be easily isolated from the reaction media. In the case of cationic mononuclear-gold(i) complexes 11, a ligand scrambling reaction in which the chloride ligand is replaced by a phosphine in the absence of a scavenger, a process previously described for gold(i) species, can be used to justify the reaction outcome.¹⁷ The formation of dinuclear gold complex 12 can be ascribed to the combination of a strong aurophilic interaction between the two gold centers (Au–Au = 2.8874(4) Å) and the stabilizing η²-coordination of the metal center to the aromatic ring of biphenylene. Similar η²-coordinated gold(i) complexes have been reported but, to the best of our knowledge, only as mononuclear species.¹⁸Taking into consideration the observed geometry of complexes 7a–b in the solid state,¹³ the facile formation of stable cationic species 11 and 12 and the lack of reactivity of the gold(i) iodides 8a–b, we hypothesized that the free rotation around the C–P bond was probably restricted, placing the gold(i) center away from the biphenylene system and thus preventing the desired oxidative insertion reaction. To overcome this problem, we set out to elongate the arm bearing the phosphine unit with an additional methylene group, introduced via a Wittig reaction from compound 4 to yield ligand 6, prepared in 4 steps in 27% overall yield. Coordination with Me₂SAuCl and AuI resulted in gold(i) complexes 9 and 10, respectively (Scheme 3). The structure of 9 was unambiguously assigned by X-ray diffraction analysis and a similar environment around the metal center to that determined for complex 7a was observed for this complex.¹³With complexes 9 and 10 in hand, we explored their reactivity towards C–C activation of the four-membered ring of biphenylene.¹⁹ After chloride abstraction and upon heating at 100 °C for 5 hours, ring opening of the biphenylene system was observed for complex 9. Interestingly, formation of mono-cyclometalated adduct 13 was exclusively observed (the structure of 13 was confirmed by ¹H, ¹³C, ³¹P, ¹⁹F, ¹¹B and 2D NMR spectroscopy and HR-MS).¹³ The solvent appears to play a major role in this process, as performing the reaction in non-chlorinated solvents resulted in stable cationic complexes similar to 11.^13,20,21 The presence of adventitious water is likely responsible for the formation of the monocyclometalated (P^C)gold(iii) complex 13 as when the reaction was carried out in C₂H₄Cl₂ previously treated with D₂O, the corresponding deuterated adduct 13-d could be detected in the reaction media. These results showcase the difficulties associated with the biscyclometalation for P-based complexes as well as the labile nature of the expected biscyclometalated adducts. Interestingly though, these processes can be seen as a de novo entry towards relatively underexplored (P^C)gold(iii) species.²²The C–C activation was further confirmed by X-ray diffraction analysis of the phosphonium salt 14, which arise from the reductive elimination at the gold(iii) center in 13 upon exchange of the BF₄⁻ counter-anion with the weakly coordinating sodium tetrakis[3,5-bis(trifluoromethyl)phenyl]borate (NaBAr^F).^13,23 The phosphorus atom is four-coordinate, with weak bonding observed to the distant counter-anion and a distorted tetrahedral geometry (C1–P–C2 = 95.05(17), C2–P–C3 = 112.1(1), C3–P–C4 = 116.6(1), C4–P–C1 = 107.4(2) deg). These results represent the third example in which the C(sp²)–P bond reductive elimination at gold(iii) has been reported.²⁴Further, it is important to note that, in contrast to the reactivity observed for the pyridine-substituted biphenylene, neither P-coordinated gold(i) iodo complexes 8a, 8b nor 10 reacted to give cyclometalated products despite prolonged heating, which highlights the need for highly reactive cationized gold(i) species to undergo oxidative addition when phosphine ligands are flanking the C–C bond.¹³To get a deeper understanding on the observed differences in reactivity for the N- vs. P-based directing groups, ground- and transition-state structures for the oxidative insertion of gold(i) halides in C1-substituted biphenylenes were computed by DFT calculations. The reactions of Py-substituted 2 with AuI to give 3 (I) and those of P-substituted 7a (II) and 9 (III) featuring the cationization of the gold(i) species were chosen as models for comparative purposes with the experimental conditions (Fig. 1 and S1–S10 in the ESI^†).^25–27 The computed activation energies for the three processes are in good agreement with the experimental data. The pyridine-substituted biphenylene I exhibits the lowest activation barrier for the oxidative insertion process (ΔG^‡ = 34.4 kcal mol⁻¹). The reaction on the phosphine-substituted derivatives II and III proved to be, after cationization of the corresponding gold(i) halide complexes (II-BF₄, III-BF₄) higher in energy (ΔG^‡ = 39.6 and 46.3 kcal mol⁻¹ respectively), although the obtained values do not rule out the feasibility of the C–C activation process. The transition state between I and I′ exhibits several interesting geometrical features: (a) the biphenylene is significantly bent, (b) the cleavage of the C–C bond is well advanced (d_C–C = 1.898 Å in TSIvs. d_C–C = 1.504 Å in I), and (c) the two C and the I atoms form a Y-shape around gold with minimal coordination from the pyridine (d_N–Au = 2.742 Å in TSIvs. d_N–Au = 2.093 Å in I and 2.157 Å in I′, respectively). The transition-state structures found for the P-based ligands (TSII and TSIII) also show an elongation of the C–C bond and display a bent biphenylene. However, much shorter P–Au distances (d_P–Au = 2.330 Å for TSII and 2.314 Å for TSIII) can be observed compared to the pyridine-based system, as expected due to the steric and electronic differences between these two coordinating groups. Analogously, longer C–Au distances were also found for the P-based systems (d_C1–Au = 2.152 Å for TSIvs. 2.235 Å and 2.204 Å for TSII and TSIII; d_C2–Au = 2.143 Å for TSIvs. 2.219 Å and 2.162 Å for TSII and TSIII), with a larger deviation of square planarity for Au in TSIII compared to TSII.^28,29 These results suggest that, provided the appropriate distance to the C–C bond is in place, the strong coordination of phosphorous to the gold(i) center does not prevent the C–C activation of biphenylene but other reactions (i.e. formation of diphosphine gold(i) cationic species, protodemetalation) can outcompete the expected biscyclometalation process. In contrast, a weaker donor such as pyridine offers a suitable balance bringing the gold in close proximity to the C–C bond and enables both the oxidative cleavage as well as the formation of the double metalation product.Open in a separate windowFig. 1Energy profile (ΔG and ΔG^‡ in kcal mol⁻¹), optimized structures, transition states computed at the IEFPCM (toluene/1,2-dichloroethane)-B3PW91/DEF2QZVPP(Au,I)/6-31++G(d,p)(other atoms) level of theory for the C–C activation of biphenylene with gold(i) iodide from I and gold(i) cationic from II and III. Computed structures of the transition states (TSI, TSII and TSIII) and table summarizing relevant distances.     

Coherent ring-current migration of Mg-phthalocyanine probed by time-resolved X-ray circular dichroism

The coherent ring current of Mg-phthalocyanine created by a broad band UV-visible pump pulse shows variation with time, where the ring currents at the corner benzene rings, around the Mg cation and on the outer ring oscillate with different time periods and the current density migrates among these regions. The 7 pairs of E_u degenerate excited states populated upon photoexcitation, generate 21 distinct coherent ring currents. We further calculate the time-resolved X-ray circular dichroism (TRXCD) spectrum of the coherences contributing to the ring current obtained by an attosecond X-ray probe pulse resonant with the nitrogen K-edge. A frequency domain TRXCD signal obtained by a Fourier transform of the signal with respect to the pump-probe delay time clearly separates the currents induced by different state pairs.

The coherent ring current of Mg-phthalocyanine are created by a broad band UV-visible pump pulse and migrate into different regions within the molecule. This coherent ring current dynamics is probed by time-resolved X-ray circular dichroism.

Controlling and measuring attosecond movements of electrons in molecules are of fundamental importance for photoinduced chemical reactions. Ring current, a circular electron motion, can be induced in cyclic conjugated molecules by applying a static magnetic field,^1–10 by using circularly polarized light (CPL)¹¹ or by coupling the molecule to an optical chiral cavity.¹² Magnetic-field-induced ring currents are widely applied in nuclear magnetic resonance,^8,13–15 where a static field non-resonant with electronic excited states perturbs the ground state and creates a ground state ring current. The direction of the magnetic field induced current is determined by the direction of the applied magnetic field and the aromaticity of the molecule, where aromatic (antiaromatic) molecules produce diamagnetic (paramagnetic) ring currents.^1,2,16–19In this theoretical study, we focus on the CPL induced ring currents, which are receiving considerable attention.^11,20–34 The sense of the resulting ring current is determined by the circular polarization of the pulse. Furthermore, CPL can create ring currents in selected states by tuning the pulse frequency to specific electronic states. This offers an additional control over the ring current generation.We consider a molecule with two excited states with perpendicular transition dipoles to the ground state. We denote the excited states |E_x〉 and |E_y〉, whose transition dipole 〈g|μ|E_x,y〉 are along the x and y direction respectively. If the bandwidth of the circularly polarized pulse with polarization covers both states, the pulse will excite them with a phase difference of , giving the electron oscillation in x and y direction a phase difference, thus driving the electrons to move in a circle.¹¹If this pair of excited states is degenerate, by a linear combination, we can create a pair of degenerate current-carrying eigenstates .^11,30 A circularly polarized ε_± light pulse increases the population of state |E_±〉 respectively. Thus, the ring current created by the pulse can be viewed as population ring current which remains time-independent after the pump pulse is over. If the excited state pair is non-degenerate, the ring current is proportional to their coherence and is denoted coherent ring current.^24,25,35 After the pulse is over, the population is stationary while the coherent ring current oscillates with a period inversely proportional to the energy difference between the coherent states.Previous research on coherent ring currents had focused on a single pair of coherence, where the current of the entire molecule evolves with the same pace.^{11,20–26,28–30} Here, we study the ring-current dynamics resulting from the superposition of multiple ring-current pairs created by a broad band circularly pump pulse. Due to the interplay of multiple coherences, we find that the evolution of ring current at different locations in the molecule show distinct feature. As the coherent current density evolves with time, we see the redistribution of current density between different regions of the molecule, which can be viewed as current migration. This behavior is analogous to the light-induced charge migration and the following evolution of electrons.^36,37,54 Localized ring currents created at different regions within the molecule, create new possibilities for designing molecular device.³⁸We focus on phthalocyanine which has broad applications in e.g. dye-sensitized solar cells^39,40 and photosensitizers for cancer photodynamic therapy.⁴¹ Its derivative Mg-phthalocyanine (MgPc) has a rigid structure (see Fig. 1), which remains stable during the electronic dynamics.^42,43 MgPc has a high density of states belonging to the E_u irreducible representation (Table S1^†), which facilitates the coherent excitation of many excited states. The molecule has multiple rings, including a central ring and the four benzene rings in the corners, which makes it possible to observe the dynamics of both local and global ring currents. We use a broadband UV pump pulse with 2 eV bandwidth made possible by recently developed attosecond UV-visible pulses.⁴⁴ Upon excitation, 7 pairs of degenerate excited states are populated within the pulse bandwidth to create 21 coherence pairs. These coherent ring currents have different profiles, giving rise to distinct evolution of local and the global ring currents. We further show that these coherences can be observed in time-resolved X-ray circular dichroism (TRXCD) spectrum resonant to the nitrogen K-edge. Contributions from each ring-current pair can be clearly identified in the Fourier-transformed CD spectrum.Open in a separate windowFig. 1(a) Sketch of the pump probe experiment. Mg-Pc is pumped by a circularly polarized UV-visible short pulse (orange arrow) and creates a coherent ring current. The coherence is detected by the TRXCD signal using an X-ray probe pulse (blue arrow) with a time delay T. (b) Loop diagram of X-ray TRXCD. Only the excited state absorption contribution is relevant to our study.As illustrated in Fig. 1, a circularly polarized pump pulse propagating along z perpendicular to the molecular x–y plane with an amplitude of 6.9934 × 10⁹ V m⁻¹, central frequency Ω = 4.0 eV and duration of τ_pu = 4.64 fs is applied at 0 fs to generate a ring current. The pulse duration and amplitude are chosen to excite 91.5% of the ground state population to excited states (Fig. S2^†), thus create a high excited state population in this multi-level system. The mechanism is similar to Rabi-flop of two-level system. The electronic wave function is obtained by numerically solving the time-dependent Schrödinger equation , where H_mol is the molecular Hamiltonian and the wave function is expanded in the molecular ground and valence excited states basis . Atomic units are used throughout. MgPc belongs to the D_4h point group. Its excited states are obtained by a TDDFT calculation. The molecular geometry is optimized using DFT with the B3LYP functional^45–48 and the 6-31g(d) basis set and Gaussian 16.⁴⁹ Both valence and core excited states are computed with TDDFT in the Tamm–Dancoff approximation (TDA) using the Chronus Quantum software package.⁵⁰ The TDA calculations employ the same density functional and basis set used in the geometry optimization. The valence excited states are listed in Table S1.^† For CPL with circularly polarization in x–y plane, the excited states of E_u irreducible representation with two-fold degeneracy are dipole allowed, and denoted mE_ux and mE_uy, where x, y denote the transition dipole direction, and m, n = 1, 2, …, 7 is the index of pair of degenerate states. The state with A_2u irreducible representation has transition dipole 〈g|μ|e〉 along z and is thus not excited by the pump pulse. The current density is given by⁵¹12where A, B run over the ground and valence excited states, is the density matrix at time t, a_B(t) is the coefficient of electronic states. D^BA_νμ is the transition one-electron density matrix (1PDM) between electronic state B and A in the atomic orbital basis and νμ denote atomic orbital. χ_μ(r) and ∇χ_μ(r) are the atomic orbital basis and its gradient respectively.The induced ring current is further probed by the TRXCD signal at the nitrogen K-edge. The Gaussian X-ray probe pulse has central frequency ω_c = 383.172 eV, σ = 0.7 fs and time delay T scanned from 0 to 25 fs. The TRXCD signal is derived in ESI S1^† using time dependent perturbation theory in the probe pulse–molecule interaction3where A, D run over the ground and valence excited states and C denotes core excited states. We employed the minimal coupling Hamiltonian for calculating the CD signal, where the transition current density is used in the CD expression instead of multipole moments.^51,52 The minimal coupling expression includes implicitly the contributions to all orders of multipolar moments.^51,52 Since CPL induces ring current only for oriented molecule,²⁰ the TRXCD signal is simulated for oriented molecules. ε_κ is the propagation unit vector of the probe pulse, and . is the momentum space transition current density. ω_DC is the transition energy from state C to D. ω is the angular frequency. Γ = 0.093 eV is core line width for nitrogen.⁵³ Here we focus on the nitrogen K-edge since the Mg line width (Γ = 0.45 eV)⁵³ is too broad to resolve different excited states as shown in Fig. S2.^†     

Correction: Mass spectrometric detection of fleeting neutral intermediates generated in electrochemical reactions

Correction for ‘Mass spectrometric detection of fleeting neutral intermediates generated in electrochemical reactions’ by Jilin Liu et al., Chem. Sci., 2021, 12, 9494–9499, DOI: 10.1039/D1SC01385H.

The authors regret that the details for ref. 15 and 17 were inadvertently swapped in the original article. The correct versions of ref. 15 and 17 are given below as ref. 1 and 2, respectively.The Royal Society of Chemistry apologises for these errors and any consequent inconvenience to authors and readers.     

Electrification of a Milstein-type catalyst for alcohol reformation

Novel energy and atom efficiency processes will be keys to develop the sustainable chemical industry of the future. Electrification could play an important role, by allowing to fine-tune energy input and using the ideal redox agent: the electron. Here we demonstrate that a commercially available Milstein ruthenium catalyst (1) can be used to promote the electrochemical oxidation of ethanol to ethyl acetate and acetate, thus demonstrating the four electron oxidation under preparative conditions. Cyclic voltammetry and DFT-calculations are used to devise a possible catalytic cycle based on a thermal chemical step generating the key hydride intermediate. Successful electrification of Milstein-type catalysts opens a pathway to use alcohols as a renewable feedstock for the generation of esters and other key building blocks in organic chemistry, thus contributing to increase energy efficiency in organic redox chemistry.

Electrification of the Milstein catalyst enabled successful molecular electrocatalytic oxidation of ethanol to the four-electron products acetate and ethyl acetate.

In order to achieve the goals of the Sustainable Development Scenario (SDS) of the International Energy Agency, the chemical industry''s emission should decline by around 10% before 2030.^1,2 This could be achieved by increasing energy efficiency and the usage of renewable feedstocks. In this respect, molecular electrocatalytic alcohol oxidation could be powerful tool by potentially providing energy and atom efficiency for organic synthesis and energy applications.^2–7 Besides the use of aminoxyl-derivatives,^8–13 especially the seminal work of Vizza, Bianchini and Grützmacher demonstrated that (transfer)-hydrogenation (TH) catalysts could be activated electrochemically and used in a so-called “organometallic fuel cell”.¹⁴ Other TH systems are however mostly limited to two electron oxidations of secondary or benzylic alcohols (Scheme 1A).^15–21Open in a separate windowScheme 1(A) Advantages/limitation of electrochemical homogeneous alcohol oxidation using well-defined catalysts. (B) Current efforts to electrify acceptor-less alcohol dehydrogenation (AAD) systems due to their large range of application in thermal catalysis.As an exception, Waymouth et al. recently reported an example of the intramolecular coupling of vicinal benzylic alcohols to the corresponding esters.^19,22 In order to extend the range of possible catalysts candidates, the Waymouth group recently also explored the possibility to use an iron-based acceptor-less alcohol dehydrogenation (AAD) catalysts²³ for electrocatalytic alcohol oxidation (Scheme 1B).²⁴ The stability under electrochemical conditions in this case is limited to <2 turnovers, but it opens the door to explore a wide range of AAD reactions under electrochemical conditions. Here, we demonstrate that a commercially available Milstein-type AAD catalyst (1)²⁵ is competent for the electrocatalytic alcohol oxidation of ethanol to ethyl acetate and acetate (Scheme 1B).The cyclic voltammogram (CV) of complex 1 (Fig. 1) shows a quasi-reversible diffusive one electron oxidation wave at 0.2 V (all potentials are referenced vs. Fc⁺/Fc⁰) in 0.2 M NaPF₆ THF/DFB (2 : 1) (DFB = 1,2 difluoro benzene) assigned to the Ru(ii)–Ru(iii) couple (see ESI, section 2.2^†). The addition of 1 to a 10 mM sodium ethoxide (NaOEt) solution in 200 mM ethanol (EtOH) in 0.1 M NaPF₆ (2 : 1 THF/DFB) gives rise to several waves at ca. −0.5, 0.0 and 0.2 V with currents significantly higher than in the absence of catalysts or substrate, indicative of possible catalytic turnover (Fig. 2). Gradual increase of the EtOH concentration from 200 mM to 1 M is accompanied by the disappearance of the first wave at −0.5 V, while a new oxidation wave appears at ca. −0.25 V (Fig. 2, light to dark green traces).Open in a separate windowFig. 1Scan rate dependence of a 1 mM solution of 1 in in 2 : 1 THF/DFB + 0.2 M NaPF6 (from light to dark green: 0.05, 0.1, 0.2, 0.3, 0.4 and 0.5 V s⁻¹, 3 mm GC electrode). Inset: evolution of the peak current as a function of the square root of the scan rate.Open in a separate windowFig. 2CVs of 10 mM NaOEt (grey) and of 5 mM 1 + 5 mM NaOEt with increasing concentrations of EtOH (from light to dark green: 200, 400, 600, 800 and 1000 mM) in 2 : 1 THF/DFB + 0.2 M NaPF₆. Scan rate 0.1 V s⁻¹, electrode: 3 mm diameter GC electrode.Increasing the base loading gradually from 5 to 20 mM yields a stark increase of current at this new wave at ca. −0.25 V (Fig. 3). Using (TBA)PF₆ instead of NaPF₆ (used to avoid Hofmann-elimination²⁶) gave similar results (see ESI, section 2.2–2.5 and section 4^†). In order to assess catalytic turnover under preparative conditions, controlled potential electrolysis (CPE) was performed. CPE experiments were run in pure ethanol (to reduce cell resistance) in the presence of 0.1 M electrolyte of well soluble bases (e.g. NaOEt, LiOH, see ESI section 4^†). CPE in 0.1 M LiOH with 1 mM 1 at E = 0 V vs. Fc^0/+ delivered ca. 15 mM of acetate and 6 mM of ethyl acetate, corresponding to 21 turnovers (per 4 electrons, or 42 turnovers per two electrons) and a faradaic efficiency (FE) of ca. 62% (see ESI section 4.3^†). In the absence of applied potential (OCP, open circuit potential), no ethyl acetate was formed (see ESI, section 4.4^†). Likewise, in the absence of catalyst, the passed charge was significantly lower (7C vs. 40C) with no detected formation of ethyl acetate. The low FE could be due to catalyst degradation, as Ru-nanoparticle formation is observed on the electrode post CPE (confirmed by SEM/Elemental mapping, see ESI section 5^†). Noteworthy, rinse-test CPE and a CPE using a simple Ru-precursor, RuCl₃, did not show any ethyl acetate formation and gave similar results to blank experiments, indicating that Ru-nanoparticles are probably not the active catalyst species and that catalyst instability could be responsible for low FE. Further studies are underway to fully understand catalyst speciation under preparative conditions (see ESI section 4.7^†) the observed catalytic activity of 1 compares well in terms of TON and product selectivity with other molecular homogeneous TH systems, with most systems being limited to the two-electron oxidation of secondary or benzylic alcohols. The Waymouth group reported a NNC ruthenium pincer for the oxidation of isopropanol to acetone with a TON of 4.¹⁸ The same group reported on the usage of phenoxy mediators with an iridium pincer complex, reaching a TON of 8 for the same reaction.²² Bonitatibus and co-workers demonstrated the activity of an iridium-based systems with a TON of 32 for the formation of p-benzaldehyde.¹⁷ Appel and co-workers reported on a nickel (TON = 3.1)¹⁵ and a cobalt triphos systems (TON = 19.9)¹⁶ for benzaldehyde formation from benzyl alcohol. To the best of our knowledge, there is only one acceptor-less alcohol dehydrogenation (AAD) catalyst that has been activated electrochemically so-far,²⁴ generating acetone with a TON <2. Only a handful of molecular systems are known to catalyze the electrochemical four electron alcohol reformation to esters, however at significantly higher potentials (1.15 V vs. Fc⁺/Fc⁰).^2,27,28 Thus, although not designed for electrochemical applications, 1 shows high activity for the challenging 4 electron oxidation of aliphatic substrates.Open in a separate windowFig. 3CV of 5 mM NaOEt (grey), 5 mM of 1 + 1 M EtOH with varying concentrations of base (5, 10, 15, and 20 mM NaOEt, light to dark green) in 2 : 1 THF/DFB + 0.2 M NaPF₆. Scan rate 0.1 V s⁻¹, electrode: 3 mm diameter GC electrode.To achieve the transposition from thermal to electrochemical TH, both Grützmacher et al. and Waymouth took advantage of a fast equilibrium between the alcohol substrate and a metal hydride intermediate that could be readily oxidized. The chemistry of ruthenium pincer AAD systems is well studied (Scheme 2)^25,29–33 and allows for a putative assignment of the observed CV-behavior. In the presence of excess base and alcohol (Fig. 2 and ​and3),3), 1 is expected to yield dearomatized complex 2,²⁵ as well as the alkoxide species 3.^25,32 We might therefore assign the first wave at −0.5 V to the oxidation of dearomatized complex 2 and the wave around 0 V to the oxidation of the alkoxide complex 3. Indeed, independently synthesized samples of 2 and 3 (in the presence of excess ethanol) give rise to oxidation half-waves at −0.45 V and −0.1 V respectively (see ESI, section 3 and 5.2^†). This is also in agreement with the observed behavior upon increasing the alcohol concentration with the expected consumption of dearomatized species 2 and concomitant disappearance of the first oxidation wave at −0.5 V. The equilibrium between 2, 3 and 4 has been reported³² and addition of excess ethanol to 2 is thus not only generating 3, but also is expected to deliver 4 (Scheme 2). The appearance of a new anodic wave at ca. −0.25 V (Fig. 2) is thus attributed to the increasing formation of 4 upon addition of larger amounts of EtOH. Complex 4 is relatively unstable in solution,^25,32,33 and decomposes in the presence of electrolyte (see ESI section 3.1^†). DFT calculations were thus used to predict its oxidation potential (see ESI, section 6^†), which was in reasonable agreement with the observed wave (−0.19 V). The DFT calculations also confirmed the assignment of the other waves related to the dearomatized complex 2 (−0.33 V) and the ethoxide species 3 (−0.1 V). A more detailed mechanistic analysis remains currently hampered by the chemical instability of 4 under the employed reaction conditions, as well as difficulties to isolate 3 in the solid state (limiting kinetic measurements). DFT calculations were thus used to get a better view on possible reaction pathways (Schemes 2, ​,33 and ESI section 6.3^†). The oxidation of 4 at −0.19 V (DFT) yields the radical cation 5, with a calculated pK_a in THF of 8.2. In the presence of NaOEt, 5 should thus deprotonate readily to give radical 6, which has an extremely negative oxidation potential of −2.1 V. At the potential it is generated, 6 should thus directly be oxidized to cationic complex 7. This cationic species 7 has a calculated pK_a of 22.7 in THF, which is in good agreement with experimental data from the Saouma group on a similar system.²⁶ The high pK_a of 7 in THF also validates the need for a strong base (e.g. NaOEt) to reform dearomatized 2. Both Grützmacher and co-workers,¹⁴ as well as Waymouth²⁴ have noted that the accelerating effect during electrocatalysis stems from the oxidation of a metal hydride intermediate that is generated by fast chemical steps. In order to verify this hypothesis and to exclude an electrochemical activation of this hydride formation step, transition state barriers were computed (Scheme 3). Taking the dearomatized complex 2 as a reference point, a first step will form the alkoxide species 3 (TS₀ = 21.2 kcal mol⁻¹). Oxidizing 2 to 8 slows down the formation of the alkoxide species (T_S0^ox = 27.5 kcal mol⁻¹), most-likely due to decreased basicity of the ligand. From the alkoxide species 3 dihydride 4 is formed via a linear, charge-separated transition state TS₁ (15.7 kcal mol⁻¹). The role of such linear transition states was highlighted recently in the case of ruthenium pincer catalysis for alcohol oxidation.^34–37 In principle, it might be envisioned that the oxidation of the metal center could be an additional driving force for this hydride abstraction step. However, after oxidation, the energy span^38,39 rises by about 11 kcal mol⁻¹ (TS₁^ox = 24.7 kcal mol⁻¹). Likewise, a beta-hydride elimination via side-arm opening is not accelerated either by oxidation (TS₂^ox = 37.5 kcal mol⁻¹, see ESI section 6.4^†). It thus seems that the generation of 4 is not accelerated by electron transfer steps and relies on a thermally activated chemical step. Importantly, alkoxide solutions were shown to be excellent hydride donors electrochemically, further corroborating that under the employed basic conditions, generation of 4 from 3 should be fast.⁴⁰ Oxidation of 4 to 5 also doesn''t accelerate thermal intramolecular release of H₂ (TS_3B^ox = 37.5 kcal mol⁻¹), which is significantly higher than neutral thermal H₂-releasing states (TS_3A and TS_3B). The experimentally observed acceleration via electron-transfer is thus proposed to follow a classical ECEC mechanism initiated by the oxidation of 4 to 5 (at roughly −0.19 V (DFT)), followed by deprotonation and re-oxidation as described above, finally delivering 2 at the electrode surface. Importantly, at the electrode surface 2 and 3 should be oxidized at the employed potentials, but based on DFT-calculations, these pathways are thought to be non-productive (Scheme 3) and could explain the low catalyst life-time and degradation under electrochemical conditions.Open in a separate windowScheme 2Reactivity of pyridine-based ruthenium complexes via dearomatization/aromatization, as well as DFT-based.Open in a separate windowScheme 3DFT-calculated energy landscape for the neutral (black dotted lines and bars) and cationic surface (blue dotted lines and bars) of ethanol dehydrogenation starting from 2 or its cationic analogue 8.     

Solvent coordination to palladium can invert the selectivity of oxidative addition

Reaction solvent was previously shown to influence the selectivity of Pd/P^tBu₃-catalyzed Suzuki–Miyaura cross-couplings of chloroaryl triflates. The role of solvents has been hypothesized to relate to their polarity, whereby polar solvents stabilize anionic transition states involving [Pd(P^tBu₃)(X)]⁻ (X = anionic ligand) and nonpolar solvents do not. However, here we report detailed studies that reveal a more complicated mechanistic picture. In particular, these results suggest that the selectivity change observed in certain solvents is primarily due to solvent coordination to palladium. Polar coordinating and polar noncoordinating solvents lead to dramatically different selectivity. In coordinating solvents, preferential reaction at triflate is likely catalyzed by Pd(P^tBu₃)(solv), whereas noncoordinating solvents lead to reaction at chloride through monoligated Pd(P^tBu₃). The role of solvent coordination is supported by stoichiometric oxidative addition experiments, density functional theory (DFT) calculations, and catalytic cross-coupling studies. Additional results suggest that anionic [Pd(P^tBu₃)(X)]⁻ is also relevant to triflate selectivity in certain scenarios, particularly when halide anions are available in high concentrations.

In the presence of the bulky monophosphine P^tBu₃, palladium usually prefers to react with Ar–Cl over Ar–OTf bonds. However, strongly coordinating solvents can bind to palladium, inducing a reversal of selectivity.

Oxidative addition is a key elementary step in diverse transformations catalyzed by transition metals.¹ For instance, this step is common to traditional cross-coupling reactions, which are among the most widely used methods for small molecule synthesis. During the oxidative addition step of cross-coupling reactions, a low valent metal [usually Pd(0)] inserts into a C–X bond with concomitant oxidation of the metal by two electrons. The “X” group of the C–X bond is commonly a halogen or triflate. Despite a wealth of research into this step,^2–5 uncertainties remain about its mechanistic nuances. The mechanistic details are especially pertinent to issues of selectivity that arise when substrates contain more than one potentially reactive C–X bond.⁶One of the best-studied examples of divergent selectivity at the oxidative addition step is the case of Pd-catalyzed Suzuki couplings of chloroaryl triflates. In 2000, Fu reported that a combination of Pd(0) and P^tBu₃ in tetrahydrofuran (THF) effects selective coupling of 1 with o-tolylB(OH)₂via C–Cl cleavage, resulting in retention of the triflate substituent in the final product 2a (Scheme 1A).⁷ In contrast, the use of PCy₃ (ref. 7) or most other phosphines⁸ provides complementary selectivity (product 2b) under similar conditions. The unique selectivity imparted by P^tBu₃ was later attributed to this ligand''s ability to promote a monoligated oxidative addition transition state on account of its bulkiness.^5,8 Smaller ligands, on the other hand, favor bisligated palladium, which prefers to react at triflate. The relationship between palladium''s ligation state and chemoselectivity has been rationalized by Schoenebeck and Houk through a distortion/interaction analysis.⁵ In brief, the selectivity preference of PdL₂ is dominated by a strong interaction between the electron-rich Pd and the more electrophilic site (C–OTf). On the other hand, PdL is less electron-rich and its selectivity preference mainly relates to minimizing unfavorable distortion energy by reacting at the more easily-distorted C–Cl bond.Open in a separate windowScheme 1Seminal reports on the effects of (A) ligands and (B) solvents on the selectivity of cross-coupling of a chloroaryl triflate.^5,7,9Proutiere and Schoenebeck later discovered that replacing THF with dimethylformamide (DMF, Scheme 1B, entry 1) or acetonitrile caused a change in selectivity for the Pd/P^tBu₃ system.^9,10 In these two polar solvents, preferential reaction at triflate was observed, and P^tBu₃ no longer displayed its unique chloride selectivity. The possibility of solvent coordination to Pd was considered, as bisligated Pd(P^tBu₃)(solv) would be expected to favor reaction at triflate. However, solvent coordination was ruled out on the basis of two intriguing studies. First, DFT calculations using the functional B3LYP suggested that solvent-coordinated transition states are prohibitively high in free energy (about 16 kcal mol⁻¹ higher than the lowest-energy monoligated transition structure). Second, the same solvent effect was not observed in a Pd/P^tBu₃-catalyzed base-free Stille coupling in DMF (Scheme 1B, entry 2). Instead, the Stille coupling was reported to favor reaction at chloride despite the use of a polar solvent. This result appears inconsistent with the possibility that solvent coordination induces triflate-selectivity, as coordination of DMF to Pd should be possible in both the Stille and Suzuki conditions, if it happens at all. Instead, it was proposed that the key difference between the Suzuki and Stille conditions was the absence of coordinating anions in the latter (unlike traditional Suzuki couplings, Stille couplings do not necessarily require basic additives such as KF to promote transmetalation). Indeed, when KF or CsF was added to the Stille reaction in DMF, selectivity shifted to favor reaction at triflate (Scheme 1B, entry 3), thereby displaying the same behavior as the Suzuki coupling in this solvent. On the basis of this and the DFT studies, it was proposed that polar solvents induce a switch in chemoselectivity if coordinating anions like fluoride are available by stabilizing anionic bisligated transition structures (Scheme 1B, right).However, our recent extended solvent effect studies produced confounding results.¹¹ In a Pd/P^tBu₃-catalyzed Suzuki cross-coupling of chloroaryl triflate 1, we observed no correlation between solvent polarity and chemoselectivity (Scheme 2). Although some polar solvents such as MeCN, DMF, and dimethylsulfoxide (DMSO) favor reaction at triflate, a number of other polar solvents provide the same results as nonpolar solvents by favoring reaction at chloride. For example, cross-coupling primarily takes place through C–Cl cleavage when the reaction is conducted in highly polar solvents like methanol, water, acetone, and propylene carbonate. In fact, the only solvents that promote reaction at triflate are ones that are commonly thought of as “coordinating” in the context of late transition metal chemistry.¹² These are solvents containing nitrogen, sulfur, or electron-rich oxygen lone pairs (nitriles, DMSO, and amides). The observed solvent effects were upheld for a variety of chloroaryl triflates and aryl boronic acids.¹¹Open in a separate windowScheme 2Expanded solvent effect studies in the Pd/P^tBu₃-catalyzed Suzuki coupling.¹¹We have sought to reconcile these observations with the earlier evidence⁹ against solvent coordination. Herein we report detailed mechanistic studies indicating that coordinating solvents alone are sufficient to induce the observed selectivity switch. In solvents like DMF and MeCN, stoichiometric oxidative addition is favored at C–OTf even in the absence of anionic additives. The apparent contradiction between our observations and the previously-reported DFT calculations and base-free Stille couplings is reconciled by a reevaluation of those studies. In particular, when dispersion is considered in DFT calculations, neutral solvent-coordinated transition structures involving Pd(P^tBu₃)(solv) become energetically feasible. Furthermore, we find that the selectivity analysis in the Stille couplings is convoluted by low yields, the formation of side products, and temperature effects. When these factors are disentangled, the Stille coupling in DMF displays selectivity similar to the Suzuki coupling in the same coordinating solvent. In light of these new results, anionic bisligated [Pd(P^tBu₃)(X)]⁻ does not appear to be the dominant active catalyst in nonpolar or polar solvents unless special measures are taken to increase the concentration of free halide, such as adding tetraalkylammonium halide salts or crown ethers.     

Tuning through-space interactions via the secondary coordination sphere of an artificial metalloenzyme leads to enhanced Rh(iii)-catalysis

We report computationally-guided protein engineering of monomeric streptavidin Rh(iii) artificial metalloenzyme to enhance catalysis of the enantioselective coupling of acrylamide hydroxamate esters and styrenes. Increased TON correlates with calculated distances between the Rh(iii) metal and surrounding residues, underscoring an artificial metalloenzyme''s propensity for additional control in metal-catalyzed transformations by through-space interactions.

We report computationally-guided protein engineering of monomeric streptavidin Rh(iii) artificial metalloenzyme to enhance catalysis of the enantioselective coupling of acrylamide hydroxamate esters and styrenes.

Artificial metalloenzymes (ArMs) can be made by anchoring a non-natural (metal) cofactor into a protein scaffold, with the goal of imbuing new-to-nature reactivity.¹ One of the most common ArM platforms is the biotin-tetrameric(strept)avidin (biotin-tSav) system pioneered by Whitesides and Ward.^2,3 These ArMs utilize high-affinity (up to K_D ∼10⁻¹⁴ M) interactions between tSav and biotin–metal conjugates. tSav-based ArMs have appeared in an increasing number of transition-metal catalyzed transformations.^4–6 In collaboration with the Ward group, we have previously described a tetrameric streptavidin (tSav) system containing a biotinylated Rh(iii) cofactor for the asymmetric synthesis of dihydroisoquinolones using benzhydroxamate esters and acrylate partners.⁷ Monomeric streptavidin (mSav), a streptavidin/rhizavidin hybrid designed to resist tetramerization, retains its high affinity for biotin (K_D ∼10⁻⁹ M).^8,9 We recently described the use of mSav as a new ArM,¹⁰ whose simpler topology encourages protein engineering via a site-directed mutagenesis approach.Traditional manipulation of a metal''s reactivity has been accomplished by modification of the electronic and steric properties of the bound ligands (Fig. 1a).^11,12 For example, we have documented and parsed the impact of Cp electronics and sterics on a number of Rh(iii) catalyzed transformations, by structural changes to the ligand in the primary coordination sphere of Rh.¹³ On the other hand, ArMs have traditionally been used as modifiers of a metal''s steric environment largely focusing on inducing asymmetry in the bond-forming events. Less broadly appreciated is the fact that any mutations in residues proximal to the active site may also impact the metal''s electronic properties via changes to the secondary coordination sphere (Fig. 1b), with the prospect of delivering more active catalysts for a given transformation.Open in a separate windowFig. 1Methods to modify the (a) primary and (b) secondary coordination sphere of a Rh(iii) catalyst.Previously, we described a mSav·Rh(iii) catalyst and demonstrated its use in the direct enantioselective coupling of acrylamide hydroxamate esters and styrenes.¹⁰ The reaction allows rapid access to piperidines – the most common N-heterocycle found in FDA-approved pharmaceuticals.¹⁴ One of the most interesting aspects of this reaction was our observation of a 7-fold increase in turnover number (TON) by embedding the cofactor into mSav''s active site.¹⁵ It has been a long-standing goal of ArMs to not only enable new-to-nature reactivity, but also for them to achieve the stellar kinetics of a native metalloenzyme. As these systems lack the evolutionary privilege of a natural metalloenzyme, extensive mutation of the protein scaffold may be required to find the optimal environment of the metal cofactor.Predicting the effects of specific mutations can prove very challenging, as any alterations to the protein conformation and charge distribution can impact reactivity regardless of the mutation''s distance from the active site.^16–19 In order to design a better mutant, we embarked on a collaborative experimental and computational study to define the role of the protein scaffold and how single point mutations affect reactivity. We identified two key residues that play a pivotal role in mSav·Rh(iii) ArM''s secondary coordination sphere, and have used this insight to design a more active mutant.For the purposes of this study, we focused on the mSav·Rh(iii) ArM-catalyzed coupling of methacrylamide with 4-methoxystyrene as our model reaction (Fig. 2a). Using a small model of the catalyst, the lowest energy pathway of this reaction''s proposed mechanism was generated (Fig. S9^†). The calculations were performed in Turbomole^20–32 with the M06 density functional.³³ Geometries were optimized with the def2-SVP basis set, and final electronic energies were calculated with the def2-TZVP basis set.³⁴ The conductor-like screening model (COSMO)³⁵ was used as implicit solvent with a dielectric of 80 to simulate water. These calculations predicted similar barriers for the N–H activation, the C–H activation, and the migratory insertion (differences less than 3 kcal mol⁻¹). Isotope-exchange experiments revealed that the C–H activation step is reversible, implicating the migratory insertion step as turnover-limiting.¹⁰Open in a separate windowFig. 2(a) Model transformation. (b) Snapshot of the transition state for alkene insertion illustrating key nearby residues Y112 (red), E124 (blue), and S119 (purple). (c) Computed barrier to alkene insertion in the presence and absence of phenol and acetate (shown in blue).The Cp* moiety of the Cp*^biotinRhX₂ cofactor is non-covalently localized in the active site likely due to a π–π stacking interaction with Y112 (Fig. 2b). This assignment is supported by the observation that mutant Y112A leads to lower yield and enantioselectivity.¹⁰ We hypothesized that we could further manipulate both the sterics and electronics of the Cp* moiety by either directly mutating Y112 or indirectly by mutating other residues that affect the Y112-Cp* interaction.To generate a model of mSav''s protein scaffold and active-site we used QM/DMD³⁶ – a hybrid quantum mechanics/molecular mechanics method that simulates proteins piecewise. Discrete molecular dynamics (DMD) equilibrates the entire system except for the metal and part of the substrate.³⁶ After a trajectory of ∼0.5 ns, quantum mechanics (QM) is used to optimize the metal region plus sidechains and residues immediately surrounding it. This process is repeated, providing efficient sampling of the entire protein scaffold while treating the metal environment quantum-mechanically. For this study, the migratory insertion transition state was modeled in WT by freezing the coordinates of the rhodium atom and the two carbon atoms forming a bond. For each system, five replicate simulations were run for ∼20 ns each.Residues E124 and S119 both hydrogen bond to Y112 and are in close proximity to the RhCp* catalytic site (Fig. 2b).³⁷ To estimate the electronic effects of these three residues on the reaction, an acetate ion, methanol molecule, and 4-methylphenol (p-cresol) molecule were added to a small catalyst model without constraints but initially positioned to mimic the sidechains of these residues (Fig. 2c). The migratory insertion energy barrier decreases by 2 kcal mol⁻¹ with incorporation of the three residues. However, this energy barrier decreases by an additional 3 kcal mol⁻¹ upon the deletion of the methanol molecule representing S119. Not only does this imply that these amino acid sidechains may be the primary reason for the increased activity of the protein-installed catalyst, but also suggest that a longer Y112–S119 distance is favorable, so long as no water can insert in this region and replace S119 in its H-bond with Y112. We hypothesize that the carboxylate group of E124 acts as a hydrogen bond acceptor, donating electron density to the Y112 phenol ring, which in turn donates electron density to the catalyst via π–π charge transfer. This could enhance the electron donation of the metal and decrease the energy barrier to the migratory insertion step. On the other hand, S119 acts as a hydrogen bond donor which would remove electron density from Y112 and subsequently the Rh(iii) moiety.Unfortunately, mutation of Y112 (Y112F and Y112W) results in negligible protein yields. We thus identified three flanking residues (T111, E113, H87) that may be expected to have a significant impact on Y112''s position, and one distal (T32) residue, chosen as distal mutations sometimes have significant impact (Fig. 3). Through this subset of mutants, we attempted to increase TON and establish a correlation between the Y112–Rh distance and Y112–S119 distance of the mutants and their reactivities.Open in a separate windowFig. 3Structure of mSav from two different views highlighting some of the mutated residues including their TON and enantioselectivity.We used QM/DMD to simulate a representative set of these mutants spanning a wide range of TONs measured in the experiment. The Y112–Rh and Y112–S119 distances were measured every ∼0.5 ps for every simulation. The results can be represented by a 3-dimensional plot with Y112–Rh distance on the X axis, Y112–S119 distance on the Y axis, and probability density on the Z axis (Fig. 4). We find the best correlation between TON and probability density in the conformational region where the Y112–Rh distance is the shortest and the Y112–S119 interaction is not energetically relevant.³⁸Open in a separate windowFig. 4Three-dimensional probability distributions from select mutants by simultaneous sampling of Rh–Y112 and S119–Y112 distances. Probabilities for the outlined regions are also shown.To clarify this correlation, we calculated the probability of having a Y112–S119 distance between 3.5–6 Å and a Y112–Rh distance less than 5.65 Å. This Y112–S119 distance corresponds to negligible hydrogen bonding.³⁹ Additionally, we constrained the small model catalyst shown in Fig. 5b (ref. 40) and calculated the corresponding energy barriers at different Y112–Rh distances (Fig. 5a). Since rate increases exponentially as the barrier decreases,³⁵ differences in probabilities in the region where the Y112–Rh distance is between 5.4–5.65 Å have the greatest impact on the relative TONs of our model methacrylamide styrene coupling. We conclude that mutants with increasing probability in this region provide increasing TON.Open in a separate windowFig. 5a) Theoretical dependence of migratory insertion barrier on Rh-phenol distance. (b) Small-model catalyst.Theoretically, a shorter Y112–Rh distance relative to WT would result in increased reactivity. Residue G49 is located under the Rh(iii) moiety (Fig. 6). We hypothesize that by mutating the glycine into an alanine, steric congestion would force the biotinylated Rh(iii) cofactor to shift upwards closer to the electron donating phenol side chain of residue Y112. Analyzing the critical portions of the Y112–Rh and Y112–S119 distances in tandem reveals that G49A has the highest probability density in this region (Fig. 4). Indeed, experimentally, this mutant gives 97 TON and 91% ee (Fig. 6). The combination of a short Y112–Rh distance and long Y112–S119 distance leads to an increase in reactivity. This is an approximate 3-fold improvement in the TON relative to WT. The G49A mutant serves as an experimental proof of concept that a computational analysis of an ArMs secondary coordination sphere can lead to the design of a more efficient ArM.Open in a separate windowFig. 6Snapshot of the transition state for alkene insertion highlighting the position of G49 (purple) relative to Rh. Y112 is shown in red and E124 is shown in blue.In summary, we have identified three key residues that contribute to accelerating the rate of a Rh(iii)-catalyzed reaction by electronic communication to the metal via the secondary coordination sphere. E124 hydrogen bonds to Y112 transferring electron density via π–π charge transfer, an effect that is attenuated by hydrogen bonding from S119. Optimal interaction of these residues can be described computationally by finding mutants that have multiple conformations bearing short Y112–Rh distances coupled with negligible bonding between Y112 and S119. This hypothesis was experimentally verified by a mutant that enforces a closer Y112–Rh distance leading to improved TON. This result demonstrates the use of a hypothesis-based site-directed mutagenesis of the secondary sphere residues, to optimize the metal''s electronic environment within the protein scaffold and enhance an ArM''s activity.