Improving the Catalytic Activity and Thermostability of MAS1 Lipase by Alanine Substitution

MAS1 is a lipase isolated from Streptomyces sp. strain W007 with potential application in biotechnology. Structural analysis of MAS1 lipase showed that eight amino acids with bulkier side located in the substrate-binding pocket may be involved in affecting catalytic performance. Alanine substitutions of those residues were conducted to reduce steric clash of catalyzed pocket and probe their functional roles. The k (cat)/K (m) of mutants H108A, F153A, and V233A increased to 2.3-, 2.1-, and 1.4-fold, respectively. Interestingly, the half-life (60 A degrees C) of F153A had shifted to 523 min after mutagenesis, which was fivefold enhancement toward that of MAS1 wide-type. Furthermore, higher hydrolysis ability of mutants H108A and F153A toward palm stearin of high melting temperature made them potentially applicable in oil/fat modification. Our work provided an example to obtain biocatalysts with desired catalytic behaviors by protein engineering.