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1.
Julia Oktawiec Henry Z. H. Jiang Ari B. Turkiewicz Jeffrey R. Long 《Chemical science》2021,12(45):15170
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. 相似文献
2.
Long Yang Becky Bongsuiru Jei Alexej Scheremetjew Binbin Yuan A. Claudia Stückl Lutz Ackermann 《Chemical science》2021,12(39):12971
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 Ep = −0.62 V vs. Ag/Ag+ and two oxidation events for the iodide, including an irreversible oxidation at Ep = 0.12 V vs. Ag/Ag+ and a reversible oxidation at Ep = 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 Eo,calc1/2 = −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. 相似文献
3.
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 acids7,8) 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 drugs28–30) 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 CuI-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 R10:1 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 (R1 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 (R1 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 R10:1 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 (RC/T). Adduction of a lysine site by LGs would reduce its accessibility to the ONAyne probe, and thus a higher RC/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 (RC/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 RC/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 RC/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. 相似文献
4.
We report here a sequential enantioselective reduction/C–H functionalization to install contiguous stereogenic carbon centers of benzocyclobutenols and cyclobutanols. This strategy features a practical enantioselective reduction of a ketone and a diastereospecific iridium-catalyzed C–H silylation. Further transformations have been explored, including controllable regioselective ring-opening reactions. In addition, this strategy has been utilized for the synthesis of three natural products, phyllostoxin (proposed structure), grandisol and fragranol.We report here a sequential enantioselective reduction/C–H functionalization to install contiguous stereogenic carbon centers of benzocyclobutenols and cyclobutanols.Molecules with inherent ring strain have gained considerable interest in the synthetic community.1 Among them, four-membered ring molecules have been recognized as powerful building blocks in organic synthesis.2 Driven by ring strain releasing, the reactions of carbon–carbon bond cleavage have been extensively studied in recent years.3 Meanwhile, cyclobutane motifs represent important structural units in natural product and bioactive molecules as well (Scheme 1).4 Therefore, a general and robust method to constitute four-membered ring derivatives is of great value, especially in an enantiomerically pure form.5Open in a separate windowScheme 1Representative cyclobutane-containing bioactive molecules.[2 + 2]-Cycloaddition6 and the skeleton rearrangement reaction7 are two primary methods to prepare chiral cyclobutane derivatives. Recently, the precision modification of four-membered ring skeletons to access enantioenriched cyclobutane derivatives has attracted emerging attention. Several strategies have been developed, including allylic alkylation,8 α-functionalization,9 conjugate addition10 and C–H functionalization11 of prochiral or racemic cyclobutane derivatives (Scheme 2a).12 However, the enantioselective synthesis of chiral benzocyclobutene derivatives is still underdeveloped.13 Although two efficient palladium-catalyzed C–H activation strategies have been developed by Baudoin14 and Martin15 groups via similar intermediate five-membered palladacycles, no enantioenriched benzocyclobutene derivative has been prepared by employing the above two methods. In 2017, Kawabata reported an elegant example of asymmetric intermolecular α-arylation of enantioenriched amino acid derivatives to afford benzocyclobutenones with tetrasubstituted carbon via memory of chirality (Scheme 2b).16 In 2018, Zhang reported an iridium-catalyzed asymmetric hydrogenation of α-alkylidene benzocyclobutenones in good enantioselectivities (3 examples, 83–88% ee).12c To the best of our knowledge, there is no report on enantioselective synthesis of benzocyclobutene derivatives with all-carbon quaternary centers.Open in a separate windowScheme 2Asymmetric synthesis of cyclobutanes and their derivatives. (a) Enantioselective functionalization of four-membered ring substrates. (b) Synthesis of chiral benzocyclobutenone via memory of chirality. (c) This work: sequential enantioselective reduction/C–H functionalization.In line with our continued interest in precision modification of four-membered ring skeletons,9d,10c,12a we initiated our studies on the synthesis of chiral benzocyclobutenes via enantioselective functionalization of highly strained benzocyclobutenones. It is well known that benzocyclobutene derivatives are labile to undergo a ring-opening reaction to release their inherent ring strains.17 Therefore, it is a challenging task to modify the benzocyclobutenone and preserve the four-membered ring skeleton at the same time. We envisioned that a carbonyl group directed C–H functionalization18 of the gem-dimethyl group could furnish enantioenriched α-quaternary benzocyclobutenones (Scheme 2c). This could be viewed as an alternative approach to achieve the alkylation of benzocyclobutenone, which was otherwise directly inaccessible using enolate chemistry through the unstable anti-aromatic intermediate.19 In addition, a highly regioselective C–H activation would be required to functionalize the methyl group instead of the aryl ring. Here we report our work on sequential enantioselective reduction and intramolecular C–H silylation to provide enantioenriched benzocyclobutenols and cyclobutanols with all-carbon quaternary centers. The excellent diastereoselectivity and regioselectivity of silylation were attributed to rigid structural organization of the 4/5 fused ring. Furthermore, this strategy has been utilized to accomplish the total synthesis of natural products phyllostoxin (proposed structure), grandisol and fragranol.We commenced our studies with enantioselective reduction of readily prepared dimethylbenzocyclobutenone 1a (Scheme 3).15,20 Surprisingly, enantioselective reduction of the carbonyl group of cyclobutanone derivatives received little attention. The first reduction of parent benzocyclobutenone was studied in 1996 by Kündig using chlorodiisopinocamphenylborane21 or chiral oxazaborolidines (CBS reduction),22 and only moderate enantioselectivity (44–68% ee) was obtained.23 Although copper-catalyzed asymmetric hydrosilylation of benzocyclobutenone 1a using CuCl/(R)-BINAP gave the benzocyclobutenol ent-2a in 88% ee, optimization of ligands gave no further improvement (Scheme 3a, see Tables S1–S4† for details).24 Gladly, excellent enantioselective reduction could be achieved in 94% yield and 97% ee under Noyori''s asymmetric transfer hydrogenation conditions (Scheme 3b, conditions A, RuCl[(S,S)-Tsdpen](p-cymene)).25 The product 2a showed remarkable stability and no ring-opening byproduct 2a′ was observed. The reduction of parent benzocyclobutenone was examined under conditions A, and benzocyclobutenol was obtained in 90% yield and 81% ee. Apparently, the steric influence imposed by the α-dimethyl group enhanced the enantioselectivity of the reduction. Similarly, the CBS reduction ((S)-B–Me) of benzocyclobutenone 1a gave better results compared with parent benzocyclobutenone, affording the product 2a in 86% yield and 92% ee (Scheme 3c).Open in a separate windowScheme 3Enantioselective reduction of benzocyclobutenone 1a. (a) Copper hydride reduction. (b) Ru-catalyzed asymmetric transfer hydrogenation. (c) CBS reduction.We then examined the substrate scope of the reduction reaction (26 was chosen to improve the yield and enantioselectivity. Besides, benzocyclobutenol 2g with nitro substitution could be obtained in 96% yield and 93% ee. Treatment of pyrrolidinyl substituted benzocycobutenone 1h with catalyst (S,S)-Ts-DENEB afforded desired product 2h in 49% yield and 89% ee, together with ring-opening product 2h′ (18%).Enantioselective reduction of benzocyclobutenonesa
Open in a separate windowaConditions A: 1a (0.5–2.0 mmol), RuCl[(S,S)-Tsdpen](p-cymene) (1–2 mol%), HCOOH/Et3N (5/2), rt. All results are corrected to the (S)-catalyst. The ee values were determined by HPLC analysis; see the ESI for more details.b(S,S)-Ts-DENEB (1–2 mol%) was used, rt or 60 °C.3,3-Disubstituted cyclobutanones were also explored (l-selectride gave cis-4i as a single product in 99% yield and 96% ee. The reaction of 3j gave similar results, and enantioenriched cyclobutanols cis-4j could be furnished in 78% yield and 97% ee from ent-trans-4j (98% ee) following the above oxidation–reduction procedure. The absolute configurations of 2a, ent-2j and trans-4i were unambiguously determined by single-crystal X-ray diffraction analysis of their corresponding nitrobenzoate derivative.27Enantioselective reduction of cyclobutanones 3a
Open in a separate windowaConditions B: 3a (1.0–5.0 mmol), (S)-B–Me (10 mol%), BH3·Me2S (0.6 equiv.), THF, rt.b(S)-B–Me (20 mol%), BH3·Me2S (1.0 equiv.).c(−)-Ipc2BCl (1.2 equiv.), THF, −20 °C. (−)-Ipc2BCl = (−)-diisopinocampheylchloroborane.Inspired by powerful and reliable directed C–H silylation chemistry pioneered by Hartwig,28 we envisioned that the transition-metal catalyzed intramolecular C–H silylations of the above alcohols would provide a single diastereomer owing to rigid structural organization. The challenges here are the control of regioselectivity in the cyclization step and inhibition of the ring-opening pathway. Benzocyclobutenol 2a was chosen as a model substrate to study this intramolecular C–H silylation. The transition-metal catalyst system and alkene acceptors were screened (Scheme 4, see Tables S5–S9† for details). Acceptor norbornene (nbe) derivative A gave the optimal yield in the cyclization step (63% NMR yield), and other phenanthroline ligands gave inferior results. The reaction showed remarkable regio- and diastereoselectivity; no silylation of the arene was detected.With optimal intramolecular silylation conditions in hand, sequential hydroxysilylation/C–H silylation/phenyllithium addition reaction of 2a provided desired product 5a in 56% overall yield without any obvious erosion of enantiomeric purity (