Ensemble-based enzyme design can recapitulate the effects of laboratory directed evolution in silico.

ABSTRACT: The creation of artificial enzymes is a key objective of computational protein design. Although de novo enzymes have been successfully designed, these exhibit low catalytic efficiencies, requiring directed evolution to improve activity. Here, we use room-temperature X-ray crystallography to study changes in the conformational ensemble during evolution of the designed Kemp eliminase HG3 (k_cat/K_M 146 M^-1s^-1). We observe that catalytic residues are increasingly rigidified, the active site becomes better pre-organized, and its entrance is widened. Based on these observations, we engineer HG4, an efficient biocatalyst (k_cat/K_M 103,000 M^-1s^-1) containing key first and second-shell mutations found during evolu

SUBMITTER: Broom A 

PROVIDER: S-EPMC7511930 | biostudies-literature | 2020 Sep

REPOSITORIES: biostudies-literature