On the Conformational Transition Between the Fully Folded and Locally Unfolded Substates of an Atypical 2-Cys Peroxiredoxin

DIEGO SEBASTIAN VAZQUEZ; ARI ZEIDA; GERARDO FERRER-SUETA; WILLIAM ARMANDO AGUDELO SUÁREZ; MONICA MONTES; JAVIER SANTOS

San Luis

Congreso; XLVIII Reunión Anual de la Sociedad Argentina de Biofísica; 2019

SAB

The thiol peroxidase from Escherichia coli (​EcTPx) is a peroxiredoxin that catalyses the reduction of different hydroperoxides. During the catalytic cycle of ​ EcTPx, two cysteine residues are involved, the peroxidatic cysteine (CP) that is first oxidized to sulfenic acid bythe substrate, and the resolving cysteine (C​R) which condenses with the sulfenic acid species of CP to form a disulfide bond. A native conformational change in ​ EcTPx, denominated ​ fully folded (FF) to ​ locally unfolded (LU) transition, involving a partial unfoldingof αH2 and αH3 helices (two non contiguous secondary structure elements), must occur to enable the formation of the disulfide bond since the catalytic cysteines are at 12 Å apart in the active-ready FF conformational substate. To elucidate this crucial process, themechanism of the FF➝LU and the LU➝FF transitions, the forward and back steps in the catalytic cycle, respectively, were studied using long time scale conventional molecular dynamic simulations in different oxidation and protonation states of C​ P​ . Helix-coil transitions usually involve overcoming high energy barriers, thus conventional molecular dynamics (MD) simulations become insufficient to study this kind of conformational rearrangements. To surpass this obstacle, we employ the accelerated molecular dynamics scheme, an enhanced sampling technique that extends the effective simulation time scale to long microseconds.Preliminary results suggest that the FF➝LU transition has a higher associated energy barrier than the refolding LU➝FF process in agreement with the experimental low catalytic rate constant of ​ EcTPx. Furthermore,​ in silico designed point-mutants of the αH3 helix enhanced locally unfolding events, suggesting that the native interactions in the active site of the FF native substate are not optimized for fully speed-up the conformational transition.