Simultaneous Improvement in the Thermostability and Catalytic Activity of Epoxidase Lsd18 for the Synthesis of Lasalocid A.

ABSTRACT: Enzymes used in the synthesis of natural products are potent catalysts, capable of efficient and stereoselective chemical transformations. Lsd18 catalyzes two sequential epoxidations during the biosynthesis of lasalocid A, a polyether polyketide natural product. We performed protein engineering on Lsd18 to improve its thermostability and catalytic activity. Utilizing structure-guided methods of FoldX and Rosetta-ddG, we designed 15 mutants of Lsd18. Screening of these mutants using thermal shift assay identified stabilized variants Lsd18-T189M, Lsd18-S195M, and the double mutant Lsd18-T189M-S195M. Trypsin digestion, molecular dynamic simulation, circular dichroism (CD) spectroscopy, and X-ray crystallography provided insights into the molecular basis for the improved enzyme properties. Not

SUBMITTER: Liu N 

PROVIDER: S-EPMC10706071 | biostudies-literature | 2023 Nov

REPOSITORIES: biostudies-literature