P-161 - Substrate triggered structural movements in Ohr: dihydrolipoamide accelerates the approximation of catalytic Arg towards the active site

Abstract

Ohr (Organic Hydroperoxide Resistance) are Cys-based, lypoil-dependent peroxidases with extraordinary reactivity towards organic hydroperoxides. The catalytic triad of Ohr comprises the reactive Cys61, Arg19 and Glu51 which interact themselves by a salt bridge network in a reduced form (the so-called closed state). Once oxidized, the Cys61 covalently binds Cys125 and the catalytic Arg would move away from the active site, as suggested by Oliveira et. al 2006. Here, we describe six new crystallographic structures from the opportunistic pathogen Chromobacterium violaceum (including the complex between Ohr and its biological reductant, dihydrolipoamide) that might represent new snapshots along the catalysis. Through classical mechanics (MM), steered molecular dynamics (SMD), hybrid quantum mechanics (QM-MM) and in vitro evaluations of point mutations, we present evidence supporting the role of dihydrolipoamide in triggering a structural switch from open to close states. Furthermore, the fully conserved Pro126 plays a crucial role allowing Cys disulfide formation during the oxidative step of catalysis. The unique biochemical and structural properties of Ohr enzymes further support their potential as drug targets, many of them present in socially relevant pathogenic microorganisms.