Dicarbonyl reduction by single enzyme for the preparation of chiral diols

Chiral diols are a group of key building blocks useful for preparing a variety of important chiral chemicals. While the preparation of optically pure diols is generally not an easy task in synthetic organic chemistry, three classes of enzymes, namely dicarbonyl reductase, dioxygenase and epoxide hydrolase, display remarkable ability to stereoselectively introduce two hydroxyl groups in a single-step enzymatic conversion. In this tutorial review, we pay special attention to dicarbonyl reductases that directly produce chiral diols through the bio-reduction of two carbonyl groups. The dicarbonyl reductases include diketoreductase, α-acetoxy ketone reductase and sepiapterin reductase. We present these exceptional enzymes in the context of source and properties, structure and catalytic mechanism as well as biocatalytic application. In addition to the broad substrate specificity, the excellent stereoselectivity and high catalytic efficiency of these enzymes have positioned them as valuable biocatalysts. With more sophisticated understanding of the structure-function relationship, the practical utilities of these enzymes associated with their interesting chemistry will be considerably appreciated over time. Moreover, rational redesign and molecular evolution of these unusual biocatalysts will truly enable their broader applications in the synthesis of chiral diols in the future.     

InspIRED by Nature: NADPH‐Dependent Imine Reductases (IREDs) as Catalysts for the Preparation of Chiral Amines

Imine reductases (IREDs) are NADPH‐dependent oxidoreductases that catalyse the asymmetric reduction of cyclic prochiral imines to amines, with excellent stereoselectivity. Since their discovery, stereocomplementary IREDs have been applied to the production of both (S) and (R) cyclic secondary amines, and the expansion in gene sequences recently identified has hinted at new substrate ranges that extend into acyclic imines and even suggest the possibility of asymmetric reductive amination from suitable ketone and amine precursors. Structural studies of various IREDs are beginning to reveal the complexities inherent in determining substrate range, stereoselectivity and mechanism in these enzymes, which represent a valuable emerging addition to the toolbox of available biocatalysts for chiral amine production.     

Cobalt cluster-containing carbonyl ylides for catalytic,three-component assembly of oxygen heterocycles

[reaction: see text] A dicobalt hexacarbonyl (Co(2)(CO)(6)) cluster is essential for the unusually broad dipolarophile scope and for the sense and degree of diastereoselection in a catalytic, three-component synthesis of tetrahydrofurans and dihydrofurans. Likely involving a new class of carbonyl ylide, these cycloadditions are stereospecific with respect to the dipolarophile and exhibit high diastereoselectivity and regioselectivity in most cases. Differentiation of all four positions of the tetrahydrofuran can thus be accomplished in a triply convergent manner.     

Stereoselective Synthesis of Trisubstituted Vinylboronates from Ketone Enolates Triggered by 1,3‐Metalate Rearrangement of Lithium Enolates

An unprecedented stereoselective synthesis of trisubstituted vinylboronates is reported to proceed by direct borylation of lithium ketone enolates under transition‐metal‐free conditions. The stereospecific C?O borylation of lithium enolates was triggered by a carbonyl‐induced 1,3‐metalate rearrangement via a C‐bound boron enolate. DFT calculations and control experiments revealed that the stereoselectivity is controlled by sterics. A variety of stereospecific trisubstituted vinylboronates, together with several tetrasubstituted vinylboronates, were conveniently synthesized with the newly developed methodology. Based on the transformation of stereospecific vinylboronate, a single isomer of Dienestrol was efficiently obtained.     

羰基还原酶辅酶结合域位点突变对其不对称催化性能的影响

基于同源模型的比较和分析,发现羰基还原酶SCR1辅酶结合域P124和W125位点对辅酶NADPH的结合形成了一定的空间位阻效应.通过对该位点进行小侧基氨基酸的取代突变,该酶的底物专一性和立体选择性均发生了不同程度的改变,表明该位点是酶与辅酶有效结合的关键位点,而且它与辅酶结合的空间效应进一步影响了底物结合域活性中心对不同构型的底物及其对映体产物的亲和作用.在底物专一性方面,野生型酶对2-羟基苯乙酮和2-溴苯乙酮及其衍生物等底物表现出较高的催化活性,而突变株W125A,W125G,P124A/W125A和P124G/W125G对苯乙酮及其部分衍生物和2-辛酮等底物的催化活性均有所提高.对于酶的立体选择性,部分突变株发生了转化产物对映体构型反转的现象,突变株P124A/W125A和P124G/W125G催化还原2-羟基苯乙酮和4-氯乙酰乙酸乙酯均生成了(R)-型产物.     

Reactivity of 2-Methylene-1,3-dicarbonyl Compounds. Stereoselective and Asymmetric Diels-Alder Reactions

The Diels-Alder reaction of 1,1-dicarbonylethenes 1 with dienes was investigated. The adduct of the reaction of 1, whose two carbonyl groups were different, with cyclopentadiene showed moderate stereoselectivity and this was explained by FMO theory. However, in the Lewis acid-catalyzed addition, the reaction proceeded with high stereoselectivity to give the exo adduct 3x. This might be due to steric hindrance because the benzene ring cannot orient in the plane of the conjugated system in the metal-chelated enedione 6. Applying this principle to (1'R,2'S,5'R)-5-methyl-2-(1-methyl-1-phenylethyl)cyclohexyl 2-benzoylacrylate (1d), we achieved a diastereomeric Diels-Alder reaction to afford 3x-R, whose structure was confirmed by the X-ray crystal analysis.     

Stereochemistry of 1,2-elimination reactions at the E2-E1cB interface--tert-butyl 3-tosyloxybutanoate and its thioester

Experimental data on the stereoselectivity of base-catalyzed 1,2-elimination reactions that produce conjugated carbonyl compounds are scarce in spite of the importance of these reactions in organic and biochemistry. As part of a comprehensive study in this area, we have synthesized stereospecifically-deuterated beta-tosyloxybutanoate esters and thioesters and studied the stereoselectivity of their elimination reactions under non-ion pairing conditions. With the availability of both the (2R*,3R*) and (2R*,3S*) diastereomers the innate stereoselectivity could be determined unambiguously. (1)H and (2)H NMR data show that these substrates produce 5-6% syn elimination, the usual amount for acyclic substrates undergoing E2 reactions. Contrary to earlier suggestions, activation by a carbonyl group has virtually no influence upon the stereoselectivity. Elimination of the (2R*,3R*) diastereomer of the beta-tosyloxyester and thioester produces 21-25% of the (Z)-alkene, much more than observed with a poorer beta-nucleofuge. A relatively large amount of (Z)-alkene product seems to be a good marker for an E2 pathway, in which the transition state is E1cB-like, rather than an E1cB(irrev) mechanism. Syn KIE values were higher than those for anti elimination for the esters as well as the thioesters. Experimental challenges to the synthesis of stereospecifically-deuterated beta-tosyloxyesters are discussed.     

Preparation and Evaluation of Arylazide-Substituted Pyridine Adenine Dinucleotides for Photoaffinity Labeling Experiments

Two pyridine-modified NAD'analogs, 3–(3-azido benzo-yl) pyridine adenine dinucleotide 1 and N -(3-azido-5-car-boxyl) phenyl nicotinamide adenine dinucleotide 2 have been prepared and evaluated for photoaffinity labeling experiments. The syntheses were accomplished via a mammalian NADase-catalyzed base exchange reaction. The new NAD⁺ analogs retained the carbonyl or carhox-amido functional group at the 3 position of the pyridine ring. The analog 1 is the first pyridine-modified azido derivative of NAD⁺ that has shown coenzyme activity in a stereospecific hydride transfer reaction catalyzed by a dehydrogenase. Both NAD⁺ analogs have shown potential for the study of active sites of NAD⁺-utilizing enzymes.     

Au-catalyzed tandem cyclization/[1,2]-alkyl migration reaction of epoxy alkynes: synthesis of spiropyranones

A novel gold-catalyzed tandem cyclization/[1,2]-alkyl migration process of epoxy alkynes to spiropyranones has been discovered. From this process, the construction of adjacent multiple stereocenters with a new quaternary carbon atom is achieved. The gold-catalyzed domino process is stereospecific with respect to the migrating carbon atom. A type of unusual C-C bond cleavage of epoxide systems has also been discovered, which can lead to the formation of two Z alkenes and a carbonyl functional group in one step with excellent stereoselectivity. Furthermore, this efficient domino process could be achieved in the presence of the simplest and least expensive gold catalyst [NaAuCl(4)].2H(2)O with a low catalyst loading.     

Rhodium cluster complexes containing bridging phenylgermanium ligands

The reaction of Rh(4)(CO)(12) with Ph(3)GeH at 97 degrees C has yielded the first rhodium cluster complexes containing bridging germylene and germylyne ligands: Rh(8)(CO)(12)(mu(4)-GePh)(6), 9, and Rh(3)(CO)(5)(GePh(3))(mu-GePh(2))(3)(mu(3)-GePh)(mu-H), 10. When the reaction is performed under hydrogen, the yield of 9 is increased to 42% and no 10 is formed. Compound 9 contains a cluster of eight rhodium atoms arranged in the form of a distorted cube. There are six mu(4)-GePh groups bridging each face of this distorted cube. Four of the rhodium atoms have two terminal carbonyl ligands, while the remaining four rhodium atoms have only one carbonyl ligand. Compound 10 contains a triangular cluster of three rhodium atoms with one terminal GePh(3) ligand, three bridging GePh(2) ligands, and one triply bridging GePh ligand. There is also one hydrido ligand that is believed to bridge one of the Rh-Ge bonds. Compound 9 reacted with PPhMe(2) at 25 degrees C to give the tetraphosphine derivative Rh(8)(CO)(8)(PPhMe(2))(4)(mu(4)-GePh)(6), 11. The structure of 11 is similar to 9 except that a PPhMe(2) ligand has replaced a carbonyl ligand on each the four Rh(CO)(2) groups. Compound 10 reacted with CO at 68 degrees C to give the complex Rh(3)(CO)(6)(mu-GePh(2))(3)(mu(3)-GePh), 12. Compound 12 is formed by the loss of the hydrido ligand and the terminal GePh(3) ligand from 10 and the addition of one carbonyl ligand. All compounds were fully characterized by IR, NMR, elemental, and single-crystal X-ray diffraction analyses.     

Platinum-rhodium carbonyl clusters: new structures and new types of dynamical activity

The reaction of Rh(4)(CO)(12) with Pt(PBu(t)(3))(2) in CH(2)Cl(2) at room temperature yielded three new complexes: Rh(4)(CO)(4)-(mu-CO)(4)(mu(4)-CO)(PBu(t)(3))(2)[Pt(PBu(t)(3))], 10, Rh(2)(CO)(8)[Pt(PBu(t)(3))](2)[Pt(CO)], 11, and Rh(2)(CO)(8)[Pt(PBu(t)(3))](3), 12. The reaction of Rh(4)(CO)(12) with an excess of Pt(PBu(t)(3))(2) in hexane at 68 degrees C yielded the new hexarhodium-tetraplatinum compound, Rh(6)(CO)(16)[Pt(PBu(t)(3))](4), 13, in a low yield. All four compounds were characterized by (31)P NMR and single-crystal X-ray diffraction analyses. Compound 10 contains an unsymmetrical quadruply bridging carbonyl ligand in the fold of a butterfly tetrahedral cluster of four rhodium atoms with a Pt(PBu(t)(3)) group bridging the hinge of the butterfly tetrahedron. Compound 11 contains an unsaturated trigonal bipyramidal Rh(2)Pt(3) cluster. Compound 12 is similar to 11 except the trigonal bipyramidal Rh(2)Pt(3) cluster opened by cleavage of one Pt-Rh bond due to steric interactions produced by the replacement of one of the carbonyl ligands in 11 with a tri-tert-butylphosphine ligand. Compound 12 undergoes facile dynamical rearrangements of the metal atoms in the cluster which average the three inequivalent phosphine ligands on the platinum atoms. Compound 13 contains an octahedral cluster of six rhodium atoms with four Pt(PBu(t)(3)) groups bridging edges of that octahedron.     

Biosynthesis of the Polycyclic System in the Antifungal HSAF and Analogues from Lysobacter enzymogenes

The biocontrol agent Lysobacter enzymogenes produces polycyclic tetramate macrolactams (PoTeMs), including the antifungal HSAF. To elucidate the biosynthesis of the cyclic systems, we identified eleven HSAF precursors/analogues with zero, one, two, or three rings through heterologous expression of the HSAF gene cluster. A series of combinatorial gene expression and deletion experiments showed that OX3 is the “gatekeeper” responsible for the formation of the first 5‐membered ring from lysobacterene A, OX1 and OX2 are responsible for formation of the second ring but with different selectivity, and OX4 is responsible for formation of the 6‐membered ring. In vitro experiments showed that OX4 is an NADPH‐dependent enzyme that catalyzes the reductive cyclization of 3‐dehydroxy alteramide C to form 3‐dehydroxy HSAF. Thus, the multiplicity of OX genes is the basis for the structural diversity of the HSAF family, which is the only characterized PoTeM cluster that involves four redox enzymes in the formation of the cyclic system.     

Highly stereoselective reagents for beta-keto ester reductions by genetic engineering of baker's yeast.

While whole cells of baker's yeast (Saccharomyces cerevisiae) are a convenient biocatalytic reducing agent for a wide variety of carbonyl compounds, mixtures of stereoisomeric alcohols are often observed since the organism contains a large number of reductase enzymes with overlapping substrate specificities but differing stereoselectivities. We sought to improve the performance of baker's yeast for beta-keto ester reductions by using recombinant DNA techniques to alter the levels of three enzymes known to play important roles in these reactions (fatty acid synthase, Fasp; aldo-keto reductase, Ypr1p; alpha-acetoxy ketone reductase, Gre2p). A complete set of "first-generation" yeast strains that either lack or overexpress each of these three enzymes was created and tested for improvements in stereoselective reductions of a series of beta-keto esters. On the basis of these results, multiply modified ("second-generation") strains were created that combined gene knockout and overexpression in single strains. In some cases, these additional modifications further improved the stereoselectivities of beta-keto ester reductions, thereby making several beta-hydroxy ester building blocks readily available by reactions that can be performed by nonspecialists. This work also revealed that additional yeast proteins participate in reducing beta-keto esters, and further progress using this strategy will require either additional genetic manipulations or the expression of yeast reductases in hosts that lack enzymes with overlapping substrate specificity.     

Investigation into the GC separation of enantiomers on a trifluoroacetylated cyclodextrin. Part II: Effect of derivatization on stereoselectivity towards alcohols

Two acyl and three fluoroacyl derivatives of 32 chiral alcohols have been chromatographed on a GC column coated with octakis(2,6-di-O-n-pentyl-3-O-trifluoroacetyl)- γ-cyclodextrin. Significant differences were observed between the stereoselectivity obtained for the derivatives and that for the underivatized alcohols. Of the derivatives, only the fluoroacylated compounds were separated into enantiomers. Derivatization with fluoroacyl groups reversed the elution order for at least some of the analytes. Stereoselectivity towards simple 2- and 3-hydroxy alkanes and their fluoroacyl derivatives was highest for those alcohols with a four-carbon chain attached to the stereogenic center. For longer-chain fluoroacyl derivative groups stereoselectivity was higher for the 2- and 3-hydroxy alkanes. Differences in stereoselectivity towards alcohols with a methyl-branched alkane chain and their fluoroacyl derivatives was related to the distance between the methyl group and the hydroxyl or fluoroacyl groups. Different degrees of saturation in the carbon chain resulted in differences in stereoselectivity. Thermodynamic data calculated for a number of analytes suggest that the alcohols and trifluoroacetate derivatives are interacting with the stationary phase by similar mechanisms. The stereospecific interaction appears to have a hydrogen bonding or dipole–dipole contribution and some form of steric component, depending upon the shape and/or size of the solute.     

Base-mediated stereospecific synthesis of aryloxy and amino substituted ethyl acrylates

The stereospecific synthesis of aryloxy and amino substituted E- and Z-ethyl-3-acrylates is of interest because of their potential in the polymer industry and in medicinal chemistry. During work on a copper-catalyzed cross-coupling reaction of ethyl (E)- and (Z)-3-iodoacrylates with phenols and N-heterocycles, we discovered a very simple (nonmetallic) method for the stereospecific synthesis of aryloxy and amino substituted acrylates. To study this long-standing problem on the stereoselectivity of aryloxy and amino substituted acrylates, a series of O- and N-substituted nucleophiles was allowed to react with ethyl (E)- and (Z)-3-iodoacrylates. Screening of different bases indicated that DABCO (1,4-diazabicyclo[2.2.2]octane) afforded successful conversion of ethyl (E)- and (Z)-3-iodoacrylates into aryloxy and amino substituted ethyl acrylates in a stereospecific manner. Herein are the details of this DABCO-mediated stereospecific synthesis of aryloxy and amino substituted E- or Z-acrylates.     

α-苯乙醇对映体在涂敷三苯甲酸纤维素酯手性固定相上的高效液相色谱法分离──洗脱液中醇的影响

在涂敷质量分数为15％的三苯甲酸纤维素酯手性柱上,考察了洗脱液中乙醇、正丙醇、异丙醇、正丁醇体积分数在色谱分离对映体性能方面的影响。初步认为在分离过程中,洗脱液中的醇与固定相的C＝O形成氢键作用,该作用和对映体与固定相的C＝O形成的氢键作用相竞争;洗脱液中醇的结构不同之所以会影响对映体的分离效果,与洗脱液中醇改变固定相中手性空穴的立体环境有关,醇的结构不同,造成固定相中手性空穴的立体环境不同。     

α-苯乙醇对映体在涂敷三苯甲酸纤维素酯手性固定相上的高效液相色谱法分离──洗脱液中醇的影响

 在涂敷质量分数为15％的三苯甲酸纤维素酯手性柱上,考察了洗脱液中乙醇、正丙醇、异丙醇、正丁醇体积分数在色谱分离对映体性能方面的影响。初步认为在分离过程中,洗脱液中的醇与固定相的C＝O形成氢键作用,该作用和对映体与固定相的C＝O形成的氢键作用相竞争;洗脱液中醇的结构不同之所以会影响对映体的分离效果,与洗脱液中醇改变固定相中手性空穴的立体环境有关,醇的结构不同,造成固定相中手性空穴的立体环境不同。     

Biocatalytic Properties and Structural Analysis of Phloroglucinol Reductases

Phloroglucinol reductases (PGRs) are involved in anaerobic degradation in bacteria, in which they catalyze the dearomatization of phloroglucinol into dihydrophloroglucinol. We identified three PGRs, from different bacterial species, that are members of the family of NAD(P)H‐dependent short‐chain dehydrogenases/reductases (SDRs). In addition to catalyzing the reduction of the physiological substrate, the three enzymes exhibit activity towards 2,4,6‐trihydroxybenzaldehyde, 2,4,6‐trihydroxyacetophenone, and methyl 2,4,6‐trihydroxybenzoate. Structural elucidation of PGRcl and comparison to known SDRs revealed a high degree of conservation. Several amino acid positions were identified as being conserved within the PGR subfamily and might be involved in substrate differentiation. The results enable the enzymatic dearomatization of monoaromatic phenol derivatives and provide insight into the functional diversity that may be found in families of enzymes displaying a high degree of structural homology.     

Synthesis and Binding of Mannose-Specific Synthetic Carbohydrate Receptors

Synthetic carbohydrate receptors (SCRs) that selectively recognize cell-surface glycans could be used for detection, drug delivery, or as therapeutics. Here we report the synthesis of seven new C_2h symmetric tetrapodal SCRs. The structures of these SCRs possess a conserved biaryl core, and they vary in the four heterocyclic binding groups that are linked to the biaryl core via secondary amines. Supramolecular association between these SCRs and five biologically relevant C¹-O-octyloxy glycans, α/β-glucoside ( α/β-Glc ), α/β-mannoside ( α/β-Man ), and β-galactoside ( β-Gal ), was studied by mass spectrometry, ¹H NMR titrations, and molecular modeling. These studies revealed that selectivity can be achieved in these tetrapodal SCRs by varying the heterocyclic binding group. We found that SCR017 (3-pyrrole), SCR021 (3-pyridine), and SCR022 (2-phenol) bind only to β-Glc. SCR019 (3-indole) binds only to β-Man. SCR020 (2-pyridine) binds β-Man and α-Man with a preference to the latter. SCR018 (2-indole) binds α-Man and β-Gal with a preference to the former. The glycan guests bound within their SCR hosts in one of three supramolecular geometries: center-parallel, center-perpendicular, and off-center. Many host–guest combinations formed higher stoichiometry complexes, 2:1 glycan⋅SCR or 1:2 glycan⋅SCR , where the former are driven by positive allosteric cooperativity induced by glycan–glycan contacts.     

Enzymatic reduction of ribonucleotides: biosynthesis pathway of deoxyribonucleotides

Ribonucleotide reductases are enzymes that synthesize the deoxyribonucleotides required for the replication of DNA in dividing cells. They thus have a key function for the growth of microorganisms and of all plant and animal tissues. The enzymes reduce all four purine and pyrimidine ribonucleotides (as the 5′-diphosphates or triphosphates) with direct substitution of the 2′-hydroxyl group by hydrogen. The physiological reducing agents are the mercapto groups of thioredoxins, a group of small proteins, which are regenerated from the oxidized form by NADPH-dependent thioredoxin reductases. There are two known types of ribonucleotide reductases (I and II), which catalyze hydrogen transfer with the aid of protein-bound iron ions or of 5′-deoxyadenosylcobalamin (coenzyme B₁₂); free radicals can be detected in both cases. The enzymes are regulated by effector nucleotides. There may exist a homeostatic mechanism, which guarantees the supply of DNA precursors to the cell.