Photobiocatalytic Cascades And Photo-induced Catalytic Promiscuity Of Enzymes

Photobiocatalysis has attracted researchers’interest recently due to its high reactivity and excellent stereoselectivity.Furthermore,photobiocatalytic cascades and the photo-excited promiscuous reactions of enzymes have found potential applications in organic synthesis.Therefore,the research on photobiocatalysis can broadly expand the substrate scope of enzymes,provide alternatives for the efficient and green synthesis of chiral compounds and contribute new insight into its characteristics and mechanism,which is of momentous scientific significance and application value.We demonstrated the photobiocatalytic cascades for the synthesis of enantiocomplementary secondary alcohols by combining photocatalytic oxidation and ketone reductase-catalyzed reduction.Alkyl benzenes were oxidized into ketones by the photocatalyst and their subsequent carbonyl reduction was catalyzed by a ketone reductase.For the model reaction of ethylbenzene,（R）-phenylethanol was obtained by Kt CR（a carbonyl reductase from Kluyveromyces thermotolerans）with 86%yield and99%e.e.,and Ytb E（a carbonyl reductase from Bacillus subtilis 168）provided（S）-phenylethanol with 34%yield and 99%e.e..A variety of substrates with different electrical substitutions on the benzene ring were explored,providing corresponding products with moderate to good results.Additionally,the deracemization of phenylethanol compounds was also realized by this cascade process.For the model deracemization of racemic 1-phenylethanol,the photobiocatalytic cascade gave（R）-1-phenylethanol by Kt CR with 95%yield and 99%e.e.and（S）-1-phenylethanol by Ras ADH（a carbonyl reductase from Ralstonia sp.DSMZ 6428）with 96%yield and99%e.e.,respectively.We reported a photo-induced promiscuous case of CHMO（cyclohexanone monooxygenase from Acinetobacter sp.NCIMB 9871）for the enantioselective synthesis ofα-fluoroketones via enantioselective reductive dehalogenation ofα,α-chlorofluoro-1-tetralone.Structure-guided rational design of CHMO improved the results remarkably,with 99%yield and 99:1 e.r by L144A and L435V for the model reaction.A possible reaction mechanism was proposed according to mechanistic studies and molecular simulation.We envisioned that theα,α-chlorofluoro-1-tetralone was bound to the active site of CHMO by some H-bonding interactions,forming an electron donor-acceptor（EDA）complex with the adjacent FADH^ˉ.The resultant EDA complex would be excited by visible light and underwent a facile single-electron transfer（SET）process with the subsequent loss of Cl^ˉto furnish theα-carbonyl radical.Then,the radical underwent SET with FADH·,followed by subsequent protonation,to provide an enzyme-associated product together with the quinone FAD.This methodology was expanded to a series of differently substituted substrates and provided highly desirable chiralα-fluoroketones in moderate to good results.The photocatalytic products,chiral fluoroketones,could be transformed into（1R,2S）-and（1S,2S）-vicinal fluoro alcohols by combining with enantiocomplementary ketoreductase.The photoexcited CHMO-catalyzed reductive dehalogenation was applied to the stereoselective synthesis of deuterated compounds.The lyophilized enzyme powder of the mutants of CHMO was used as the catalyst,D₂O as the solvent and deuterium source,andα,α-chlorofluoro-1-tetralone as the substrate.The influence factors of the reaction such as the types and proportions of cosolvent,temperature,enzyme amount and reaction time were investigated.The model substrate was transformed into the deuterated product with 92%yield,98%e.e.and 96%D-incorporation.With the optimized conditions in hand,a series of differently substituted substrates were transformed into the corresponding products with 58-92%yields,91-98%e.e.and 94-97%D-incorporation by L144A or L435V.It also contributed toα-deuterated 2-methyl-1-tetralone with 90%yield,98%e.e.and 93%D-incorporation,andα-deuterated donepezil with 66%yield and 70%D-incorporation,demonstrating its great potential in the synthesis of deuterium drugs.Lastly,the photo-induced Giese radical addition by Lb ADH（a carbonyl reductase from Lactobacillus brevis）was investigated using benzyl iodine and 2-methylbutyrolactone as substrates.The engineered Lb ADH（E144F/V195F）significantly improved the activity and stereoselectivity with 77%yield and 95%e.e..Furthermore,this methodology was expanded to a series of differently substituted benzyl iodine,and provided the highly desirable chiral product with 85%-93%e.e..Additionally,a possible reaction mechanism was proposed on the basis of mechanism experiments.