Heterologous expression of peptidyl-Lys metallopeptidase of Armillaria mellea and mutagenic analysis of the recombinant peptidase.

ABSTRACT: A method to express, purify and modify the Peptidyl-Lys metallopeptidase (LysN) ofArmillaria melleainPichia pastoriswas developed to enable functional studies of the protease. Based on prior work, we propose a mechanism of action of LysN. Catalytic residues were investigated by site-directed mutagenesis. As anticipated, these mutations resulted in significantly reduced catalytic rates. Additionally, based on molecular modelling eleven mutants were designed to have altered substrate specificity. The S1' binding pocket of LysN is quite narrow and lined with negative charge to specifically accommodate lysine. To allow for arginine specificity in S1', it was proposed to extend the S1' binding pocket by mutagenesis, however the resulting mutant did not show any activity with arginine in P1'. Tw

SUBMITTER: Odum AS 

PROVIDER: S-EPMC4885931 | biostudies-literature | 2016 Apr

REPOSITORIES: biostudies-literature