Do-it-yourself histidine-tagged bovine enterokinase: a handy member of the protein engineer's toolbox.

ABSTRACT: Enterokinase, a two-chain duodenal serine protease, activates trypsinogen by removing its N-terminal propeptide. Due to a clean cut after the non-primed site recognition sequence, the enterokinase light chain is frequently employed in biotechnology to separate N-terminal affinity tags from target proteins with authentic N-termini. In order to obtain large quantities of this protease, we adapted an in vitro folding protocol for a pentahistidine-tagged triple mutant of the bovine enterokinase light chain. The purified, highly active enzyme successfully processed recombinant target proteins, while the pentahistidine-tag facilitated post-cleavage removal. Hence, we conclude that producing enterokinase in one's own laboratory is an efficient alternative to the commercial enzyme.

SUBMITTER: Skala W 

PROVIDER: S-EPMC3863954 | biostudies-literature | 2013 Dec

REPOSITORIES: biostudies-literature