Generation of Oxidoreductases with Dual Alcohol Dehydrogenase and Amine Dehydrogenase Activity

The l -lysine-ϵ-dehydrogenase (LysEDH) from Geobacillus stearothermophilus naturally catalyzes the oxidative deamination of the ϵ-amino group of l -lysine. We previously engineered this enzyme to create amine dehydrogenase (AmDH) variants that possess a new hydrophobic cavity in their active site such that aromatic ketones can bind and be converted into α-chiral amines with excellent enantioselectivity. We also recently observed that LysEDH was capable of reducing aromatic aldehydes into primary alcohols. Herein, we harnessed the promiscuous alcohol dehydrogenase (ADH) activity of LysEDH to create new variants that exhibited enhanced catalytic activity for the reduction of substituted benzaldehydes and arylaliphatic aldehydes to primary alcohols. Notably, these novel engineered dehydrogenases also catalyzed the reductive amination of a variety of aldehydes and ketones with excellent enantioselectivity, thus exhibiting a dual AmDH/ADH activity. We envisioned that the catalytic bi-functionality of these enzymes could be applied for the direct conversion of alcohols into amines. As a proof-of-principle, we performed an unprecedented one-pot “hydrogen-borrowing” cascade to convert benzyl alcohol to benzylamine using a single enzyme. Conducting the same biocatalytic cascade in the presence of cofactor recycling enzymes (i.e., NADH-oxidase and formate dehydrogenase) increased the reaction yields. In summary, this work provides the first examples of enzymes showing “alcohol aminase” activity.     

Facile Access to Chiral Alcohols with Pharmaceutical Relevance Using a Ketoreductase Newly Mined from Pichia guilliermondii

Chiral secondary alcohols with additional functional groups are frequently required as important and valuable synthons for pharmaceuticals, agricultural and other fine chemicals. With the advantages of environmentally benign reaction conditions, broad reaction scope, and high stereoselectivity, biocatalytic reduction of prochiral ketones offers significant potential in the synthesis of optically active alcohols. A CmCR homologous carbonyl reductase from Pichia guilliermondii NRRL Y‐324 was successfully overexpressed. Substrate profile characterization revealed its broad substrate specificity, covering aryl ketones, aliphatic ketones and ketoesters. Furthermore, a variety of ketone substrates were asymmetrically reduced by the purified enzyme with an additionally NADPH regeneration system. The reduction system exhibited excellent enantioselectivity (>99% ee) in the reduction of all the aromatic ketones and ketoesters, except for 2‐bromoacetophenone (93.5% ee). Semi‐preparative reduction of six ketones was achieved with high enantioselectivity (>99% ee) and isolation yields (>80%) within 12 h. This study provides a useful guidance for further application of this enzyme in the asymmetric synthesis of chiral alcohol enantiomers.     

光学纯SunPhos手性双膦烷的合成及其与丙烯酸甲酯的反应

SunPhos 双膦烷可以用氢化铝锂和三甲基氯硅烷还原相应的光学纯的双磷酸酯获得，该双膦烷和丙烯酸甲酯反应，可得膦烷被双烷基化的富电子膦配体。     

Chemoenzymatic Synthesis of Enantiopure Amino Alcohols from Simple Methyl Ketones

This study highlights the straight-forward synthesis of substituted 1,2-amino alcohols from simple and readily available aromatic methyl ketones. Starting from acetophenone derivatives, the straight-forward synthesis strategy involved an initial bromination of the alpha-positioned methyl group in the first step, followed by a simple hydrolysis to the hydroxyketone (2-hydroxyacetophenone). The hydroxylated intermediate was subsequently converted from Silicibacter pomeroyi to the final 1,2-amino alcohol by using the transaminase. The transaminase-catalyzed reaction proceeded with yields up to 62 % and always excellent enantiomeric excess of >99 %. Interestingly, the keto-enol-tautomerism of the hydroxyl ketone yields an unexpected amino alcohol isomer.     

Pyrroloquinoline Quinone Aza-Crown Ether Complexes as Biomimetics for Lanthanide and Calcium Dependent Alcohol Dehydrogenases**

Understanding the role of metal ions in biology can lead to the development of new catalysts for several industrially important transformations. Lanthanides are the most recent group of metal ions that have been shown to be important in biology, that is, in quinone-dependent methanol dehydrogenases (MDH). Here we evaluate a literature-known pyrroloquinoline quinone (PQQ) and 1-aza-15-crown-5 based ligand platform as scaffold for Ca²⁺, Ba²⁺, La³⁺ and Lu³⁺ biomimetics of MDH and we evaluate the importance of ligand design, charge, size, counterions and base for the alcohol oxidation reaction using NMR spectroscopy. In addition, we report a new straightforward synthetic route (3 steps instead of 11 and 33 % instead of 0.6 % yield) for biomimetic ligands based on PQQ. We show that when studying biomimetics for MDH, larger metal ions and those with lower charge in this case promote the dehydrogenation reaction more effectively and that this is likely an effect of the ligand design which must be considered when studying biomimetics. To gain more information on the structures and impact of counterions of the complexes, we performed collision induced dissociation (CID) experiments and observe that the nitrates are more tightly bound than the triflates. To resolve the structure of the complexes in the gas phase we combined DFT-calculations and ion mobility measurements (IMS). Furthermore, we characterized the obtained complexes and reaction mixtures using Electron Paramagnetic Resonance (EPR) spectroscopy and show the presence of a small amount of quinone-based radical.     

Synthesis of Novel Chiral Ionic Liquid and Its Application in Reduction of Prochiral Ketones to the Corresponding Chiral Alcohols Using NaBH4

A novel chiral ionic liquid (CIL) based on nicotinium salt has been synthesized and used as an efficient asymmetric chiral catalyst for reduction of acetophenone derivatives with NaBH₄ in methanol at room temperature. The optically active alcohols were obtained in low to moderate enantiomeric excess in a short reaction time.     

Facile Synthesis of Enantiopure Sugar Alcohols: Asymmetric Hydrogenation and Dynamic Kinetic Resolution Combined

An unprecedented Ir/f‐amphox‐catalyzed asymmetric hydrogenation of racemic 2,3‐syn‐dihydroxy‐1,4‐diones is presented involving dynamic kinetic resolution, which produces (1R,2R,3R,4R)‐tetraols. This protocol constitutes an efficient and straightforward approach to accessing sugar alcohols bearing four contiguous stereocenters. The strategy exhibits various advantages over existing methods, including excellent yields (up to 98 %), exceptional stereoselectivities (up to 99:1 dr, 99.9 % ee), operational simplicity and substrate generality. Moreover, the nature of the reaction was revealed as a stepwise transformation by in situ Fourier‐transform infrared spectroscopy and isolation of intermediates.     

Synthesis and Structure of an Enantiopure Dirhodium Loop with Unusual Axial Coordination

The crystals of enantiopure SS-[cis-Rh2(Ph2C6H4)2(O2CC2F4CO2)]2((CH3)2CHCH2NH2)3(1) were obtained from the reaction of S-[cis-Rh2(Ph2C6H4)2(CH3CN)6]BF4(S-2) and(Et4N)2(O2CC2F4CO2) in CH2Cl2/CH3OH under the presence of sec-butyl amine.Compound 1 crystallizes in orthorhombic,space group P212121 with a = 16.880(5),b = 28.728(9),c = 20.475(6) ,V = 9929(5) 3,Z = 4,C95H96Cl5F8N3O8.50P4Rh4,Mr = 2280.52,Dc = 1.526 g/cm3,F(000) = 4608 and μ(MoKα) = 0.922 mm-1.The final R = 0.0488 and wR = 0.1164 for 17204 observed reflections with I > 2σ(I) and R = 0.0814 and wR = 0.1374 for all data with absolute structure parameter =-0.04(3).Compound 1 contains two inherently chiral S-[cis-Rh2(Ph2C6H4)2] moieties which are connected by two(O2CC2F4CO2) dicarboxylate ligands in the equatorial positions.One of the dirhodium units with Rh-Rh distance of 2.5445(8)  is further coordinated by a sec-butyl amine in each axial position.Another dirohdium unit has only one axial sec-butyl amine ligand,and its Rh-Rh distance is 2.5079(9) .     

Catalytic Asymmetric Synthesis of Allylic Alcohols and Derivatives and their Applications in Organic Synthesis

Allylic alcohols represent an important and highly versatile class of chiral building blocks for organic synthesis. This Review summarizes the plethora of methods developed for the catalytic asymmetric synthesis of enantioenriched allylic alcohols. These include: dynamic kinetic resolution (DKR/DKAT), nucleophilic 1,2‐addition to carbonyl groups, allylic substitution, oxidation of C? H bonds, the addition of O nucleophiles to π systems, reduction of unsaturated carbonyl compounds, and an alternative route from enantioenriched propargylic alcohols. Furthermore, these catalytic asymmetric processes are exemplified by their applications in the syntheses of complex molecules such as natural products and potential therapeutic agents.     

Ruthenium‐Catalyzed Tandem‐Isomerization/Asymmetric Transfer Hydrogenation of Allylic Alcohols

A one‐pot procedure for the direct conversion of racemic allylic alcohols to enantiomerically enriched saturated alcohols is presented. The tandem‐isomerization/asymmetric transfer hydrogenation process is efficiently catalyzed by [{Ru(p‐cymene)Cl₂}₂] in combination with the α‐amino acid hydroxyamide ligand 1 , and performed under mild conditions in a mixture of ethanol and THF. The saturated alcohol products are isolated in good to excellent chemical yields and in enantiomeric excess up to 93 %.     

Diversity‐Oriented Enantioselective Synthesis of Highly Functionalized Cyclic and Bicyclic Alcohols

The copper‐catalyzed hetero‐allylic asymmetric alkylation (h‐AAA) of functionalized Grignard reagents that contain alkene or alkyne moieties has been achieved with excellent regio‐ and enantioselectivity. The corresponding alkylation products were further transformed into a variety of highly functionalized cyclic and bicyclic alcohols with excellent control over the chemo‐, regio‐, and stereoselectivity.     

2,3,5,6-四氟苯甲醇的合成

经氟代、水解脱羧、酯化、还原等步骤合成了杀虫剂四氟苯菊酯的重要中间体2,3,5,6-四氟苯甲醇, 改进了氟代反应的无水操作和反应条件, 产物四氟对苯二甲腈纯度高达98.3%, 通过加入水参与反应改进了水解反应, 使水解和脱羧由两步反应变为一步, 且产物为只脱一个羧基的2,3,5,6-四氟苯甲酸, 收率可以高达92.5%, 用相对价廉的NaBH₄/I₂体系还原2,3,5,6-四氟苯甲酸甲酯以52.3%的收率得到了目标产物, 总收率29.6%.     

Catalyst‐Free Dehydrative α‐Alkylation of Ketones with Alcohols: Green and Selective Autocatalyzed Synthesis of Alcohols and Ketones

Direct dehydrative α‐alkylation reactions of ketones with alcohols are now realized under simple, practical, and green conditions without using external catalysts. These catalyst‐free autocatalyzed alkylation methods can efficiently afford useful alkylated ketone or alcohol products in a one‐pot manner and on a large scale by C?C bond formation of the in situ generated intermediates with subsequent controllable and selective Meerwein–Pondorf–Verley–Oppenauer‐type redox processes.     

Fluorometric Assay of Alcohols Using Alcohol Oxidase

Abstract

Fluorometric methods are described for the assay of ethanol, methanol, allyl alcohol and n-propanol using the enzyme alcohol oxidase. The enzyme catalyzes the oxidation of the alcohols to aldehydes with concomitant formation of hydrogen peroxide. The hydrogen peroxide formed is measured fluorometrically using p-hydroxy-phenylacetic acid and peroxidase. From 0.1–100 μg/ml of the alcohols and from 0.0002 to 0.050 unit/ml of alcohol oxidase can be assayed in 2 minutes with a precision of about ± 2%.     

One‐Pot Cascade Biotransformation for Efficient Synthesis of Benzyl Alcohol and Its Analogs

Benzyl alcohol is a naturally occurring aromatic alcohol and has been widely used in the cosmetics and flavor/fragrance industries. The whole‐cell biotransformation for synthesis of benzyl alcohol directly from bio‐based L‐phenylalanine (L‐Phe) was herein explored using an artificial enzyme cascade in Escherichia coli. Benzaldehyde was first produced from L‐Phe via four heterologous enzymatic steps that comprises L‐amino acid deaminase (LAAD), hydroxymandelate synthase (HmaS), (S)‐mandelate dehydrogenase (SMDH) and benzoylformate decarboxylase (BFD). The subsequent reduction of benzaldehyde to benzyl alcohol was achieved by a broad substrate specificity phenylacetaldehyde reductase (PAR) from Solanum lycopersicum. We found the designed enzyme cascade could efficiently convert L‐Phe into benzyl alcohol with conversion above 99%. In addition, we also examined L‐tyrosine (L‐Tyr) and m‐fluoro‐phenylalanine (m‐f‐Phe) as substrates, the cascade biotransformation could also efficiently produce p‐hydroxybenzyl alcohol and m‐fluoro‐benzyl alcohol. In summary, the developed biocatalytic pathway has great potential to produce various high‐valued fine chemicals.