A bacterial display system for effective selection of protein-biotin ligase BirA variants with novel peptide specificity.

ABSTRACT: Biotinylation creates a sensitive and specific tag for purification and detection of target proteins. The E. coli protein-biotin ligase BirA biotinylates a lysine within a synthetic biotin acceptor peptide (AP) and allow for specific tagging of proteins fused to the AP. The approach is not applicable to unmodified proteins, and we sought to develop an effective selection system that could form the basis for directed evolution of novel BirA variants with specificity towards unmodified proteins. The system was based on bacterial display of a target peptide sequence, which could be biotinylated by cytosolic BirA variants before being displayed on the surface. In a model selection, the bacterial display system accomplished >1.000.000 enrichment in a single selection step. A randomly mutated Bi

SUBMITTER: Granhoj J 

PROVIDER: S-EPMC6411976 | biostudies-literature | 2019 Mar

REPOSITORIES: biostudies-literature