Catalytic promiscuity of Escherichia coli BioH esterase: Application in the synthesis of 3,4-dihydropyran derivatives

Abstract

Enzymatic catalytic promiscuity has received increasing attention in the past decade. In this research, ten enzymes were investigated for the promiscuous activity in catalysis of the Michael addition-cyclization cascade reaction of p-nitrobenzalacetone with 1,3-cyclohexanedione to prepare 2-hydroxy-2-methyl-4-(4-nitrophenyl)-3,4,7,8-tetrahydro-2H-chromen-5(6H)-one in anhydrous media, and control experiments were conducted to exclude false positive results. The highest yield (46.1%) was observed with Escherichia coli BioH esterase and the optimal reaction condition was: 1mmol α,β-unsaturated ketone, 1mmol 1,3-dicarbonyl compound, 20mg E. coli BioH esterase, 20ml N,N-dimethylformamide at 37°C for 120h. To preliminarily investigate the mechanism, site-directed mutagenesis was performed on the hydrolysis catalytic triad of BioH, and the results indicated “alternate-site enzyme promiscuity”. When a series of substituted benzalacetones and 1,3-cyclic diketones were used as the reactants, yields of up to 76.3% were achieved. These results imply the potential industrial application of E. coli BioH in the preparation of dihydropyran derivatives.