Engineering of Ancestors as a Tool to Elucidate Structure, Mechanism, and Specificity of Extant Terpene Cyclase.

ABSTRACT: Structural information is crucial for understanding catalytic mechanisms and to guide enzyme engineering efforts of biocatalysts, such as terpene cyclases. However, low sequence similarity can impede homology modeling, and inherent protein instability presents challenges for structural studies. We hypothesized that X-ray crystallography of engineered thermostable ancestral enzymes can enable access to reliable homology models of extant biocatalysts. We have applied this concept in concert with molecular modeling and enzymatic assays to understand the structure activity relationship of spiroviolene synthase, a class I terpene cyclase, aiming to engineer its specificity. Engineering a surface patch in the reconstructed ancestor afforded a template structure for generation of a high-confidenc

SUBMITTER: Schriever K 

PROVIDER: S-EPMC8023661 | biostudies-literature | 2021 Mar

REPOSITORIES: biostudies-literature