Hydrolase-catalyzed Directed Asymmetry Aldol Reaction

Aldol addition is a kind of important classic reaction in organic synthesis which has the ability to build new types of C-C bond. Aldol addition can be naturally catalyzed by aldolases which exist in organisms. But beyond that, there are very few enzymes which could catalyze Aldol reaction, so the study of enzyme catalytic promiscuity on Aldol reaction will be of great significance.CAL-B mutants were found to catalyze asymmetric Aldol addition of cyclohexanone and aromatic aldehyde in organic media.24kinds of CAL-B mutants were obtained through error-prone PCR. The influence of reaction conditions including different kinds of CAL-B mutants, temperature, enzyme concentration, water content and the structures of substrates on CAL-B mutants-catalyzed asymmetric Aldol reaction were systematically investigated. The experiment results demonstrated that the active site of CAL-B was responsible for the enzymatic Aldol addition reaction. Among these mutants, B12was the best mutan by which we could get the excellent yield and ee with the addition of water, both were99%. Also we have examined the scope of substrates for the asymmetric Aldol reaction under CAL-B mutant B12.Meantime, amide was found to have great influnce on the asymmetric Aldol reaction catalyzed by acidic protease and alkaline protease in organic media. Aldol addition of cyclohexanone and aromatic aldehyde was explored. The influence of reaction conditions including enzyme source, types of amides, temperature, enzyme concentration and the structures of substrate on protease-catalyzed asymmetric Aldol reaction were systematically investigated. The control experiment results demonstrated that the asymmetric Aldol reaction catalyzed by acidic protease and alkaline protease can be assisted by formamide which also improves stereoselectivity of the reaction. We could get the best ee (81%,68%respectively) in the system of acidic protease/formamide and alkaline protease/formamide.In this thesis,10Aldol reaction products were synthesized. These compounds were characterized by1H NMR and analysized by chairal HPLC.