Linked production of pyroglutamate-modified proteins via self-cleavage of fusion tags with TEV protease and autonomous N-terminal cyclization with glutaminyl cyclase in vivo.

ABSTRACT: Overproduction of N-terminal pyroglutamate (pGlu)-modified proteins utilizing Escherichia coli or eukaryotic cells is a challenging work owing to the fact that the recombinant proteins need to be recovered by proteolytic removal of fusion tags to expose the N-terminal glutaminyl or glutamyl residue, which is then converted into pGlu catalyzed by the enzyme glutaminyl cyclase. Herein we describe a new method for production of N-terminal pGlu-containing proteins in vivo via intracellular self-cleavage of fusion tags by tobacco etch virus (TEV) protease and then immediate N-terminal cyclization of passenger target proteins by a bacterial glutaminyl cyclase. To combine with the sticky-end PCR cloning strategy, this design allows the gene of target proteins to be efficiently inserted into the e

SUBMITTER: Shih YP 

PROVIDER: S-EPMC3986218 | biostudies-literature | 2014

REPOSITORIES: biostudies-literature