Mutations at the putative active cavity of styrene monooxygenase: enhanced activity and reversed enantioselectivity.
Styrene monooxygenase (SMO) catalyzes the first step of styrene degradation, and also serves as an important enzyme for the synthesis of enantiopure epoxides. To enhance its activity, molecular docking of styrene was performed based on the X-ray crystal structure of the oxygenase subunit of SMO to identify three amino acid residues (Tyr73, His76 and Ser96) being adjacent to the phenyl ring of styrene. Variants at those positions were constructed and their enzymatic activities were analyzed. Three mutants (Y73V, Y73F, and S96A) were found to exhibit higher enzymatic activities than the wild-type in the epoxidation of styrene, while retaining excellent stereoselectivity. The specific epoxidation activity of the most active mutant S96A toward styrene and trans-β-methyl styrene were 2.6 and 2.3-fold of the wild-type, respectively. In addition, the Y73V mutant showed an unexpected reversal of enantiomeric preference toward 1-phenylcyclohexene.
Key Laboratory of Environmental and Applied Microbiology, Chengdu Institute of Biology, Chinese Academy of Sciences, P.O. Box 416, Chengdu 610041, PR China.
Hui
H
Lin
De-Fang
DF
Tang
Abeer Ahmed Qaed
AA
Ahmed
Yan
Y
Liu
Zhong-Liu
ZL
Wu
D000595:Amino Acid Sequence; D055162:Biocatalysis; D001711:Biotransformation; D020134:Catalytic Domain / Q000235:genetics*; D004852:Epoxy Compounds / Q000378:metabolism; D008969:Molecular Sequence Data; D050505:Mutant Proteins / Q000737:chemistry / Q000378:metabolism; D009154:Mutation / Q000235:genetics*; D010105:Oxygenases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D016958:Pseudomonas putida / Q000201:enzymology*; D016415:Sequence Alignment; D013237:Stereoisomerism; D020058:Styrene / Q000737:chemistry / Q000378:metabolism; D013379:Substrate Specificity
D004852:Epoxy Compounds; D050505:Mutant Proteins; D020058:Styrene; D010105:Oxygenases; C026721:styrene monooxygenase