Enhancing the promiscuity of a member of the Caspase protease family by rational design.

ABSTRACT: The N-terminal cleavage of fusion tags to restore the native N-terminus of recombinant proteins is a challenging task and up to today, protocols need to be optimized for different proteins individually. Within this work, we present a novel protease that was designed in-silico to yield enhanced promiscuity toward different N-terminal amino acids. Two mutations in the active-site amino acids of human Caspase-2 were determined to increase the recognition of branched amino-acids, which show only poor binding capabilities in the unmutated protease. These mutations were determined by sequential and structural comparisons of Caspase-2 and Caspase-3 and their effect was additionally predicted using free-energy calculations. The two mutants proposed in the in-silico studies were expressed and in-vi

SUBMITTER: Ohlknecht C 

PROVIDER: S-EPMC7497161 | biostudies-literature | 2020 Oct

REPOSITORIES: biostudies-literature