Tuning Enzyme Thermostability via Computationally Guided Covalent Stapling and Structural Basis of Enhanced Stabilization.

ABSTRACT: Enhancing the thermostability of enzymes without impacting their catalytic function represents an important yet challenging goal in protein engineering and biocatalysis. We recently introduced a novel method for enzyme thermostabilization that relies on the computationally guided installation of genetically encoded thioether "staples" into a protein via cysteine alkylation with the noncanonical amino acid O-2-bromoethyl tyrosine (O2beY). Here, we demonstrate the functionality of an expanded set of electrophilic amino acids featuring chloroacetamido, acrylamido, and vinylsulfonamido side-chain groups for protein stapling using this strategy. Using a myoglobin-based cyclopropanase as a model enzyme, our studies show that covalent stapling with p-chloroacetamido-phenylalanine (p

SUBMITTER: Iannuzzelli JA 

PROVIDER: S-EPMC9178789 | biostudies-literature | 2022 May

REPOSITORIES: biostudies-literature