Accessing to local unfolding events that take place during the catalytic cycle of E. coli thiol peroxidase: insights from accelerated molecular dynamic simulation profiles

DIEGO SEBASTIAN VAZQUEZ; AGUDELO, WILLIAM A.; FERRER-SUETA, GERARDO; JAVIER SANTOS

Foz do Iguaçu, Brazil

Congreso; 23rd Congress of the IUMBM and 44th Annual Meeting of the Brazilian Society for Biochemistry and Molecular Biology; 2015

International Union for Biochemistry and Molecular Biology & Brazilian Society for Biochemistry and Molecular Biology

Introduction and objectives: Peroxiredoxins are ubiquitous enzymes that reduce different kind of peroxides via a reactive cysteine. The active fully folded (FF) conformation of the thiolperoxidase (TPX) requires a key peroxidatic cysteine (CP, located in helix-α2). First, CP is oxidized tosulfenic acid (CysOH) and then, after a critically local unfolding step (LU), CP forms a disulfide bond with the resolving cysteine (C R, located in helix-α 3, Fig.1A-C). We hypothesize that (i) the helix α 3 has an intrinsic tendency to unfold and (ii) the conformational change involved is the rate-limiting step in the catalytic cycle. In order to study these events and the structural determinants of the C P-CR disulfide bond formation, we performed biased conformational searching profiles, followed by PCA analysis altogether with experimental measurement of the thermodynamic stability of FF and LU states.Materials and methods: Common and accelerated (aMD) simulations were carried out and analyzed in the AMBER14-GPU package using TIP3P waters at 300K and NVT ensemble. PCA analysis was performed to identify populations of conformations based on their dynamics.Results and conclusions: Preliminary results from aMD simulations of the FF→LU and LU→FF processes suggest that the latter has a lower transition barrier, as judged by the lower acceleration factor needed to reach the other basin.