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

ZEIDA ARI; MÓNICA R. MONTES , JOSÉ L. MONTI , RODOLFO M. GONZÁLEZ-LEBRERO, ALEJANDRO J. SPIAGGI, PATRICIO J. GARRAHAN Y ROLANDO C. ROSSI.; GERARDO FERRER-SUETA; SANTOS JAVIER; VAZQUEZ DIEGO; WILLIAM A. AGUDELO

San Luis

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

Sociedad Argentina de Biofisica

The thiol peroxidase from Escherichia coli ( Ec TPx) is a peroxiredoxin that catalysesthe reduction of different hydroperoxides. During the catalytic cycle of Ec TPx, two cysteineresidues are involved, the peroxidatic cysteine (C P ) that is first oxidized to sulfenic acid bythe substrate, and the resolving cysteine (C R ) which condenses with the sulfenic acidspecies of C P to form a disulfide bond. A native conformational change in Ec TPx,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 toenable the formation of the disulfide bond since the catalytic cysteines are at 12 Å apart inthe 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 thecatalytic cycle, respectively, were studied using long time scale conventional moleculardynamic simulations in different oxidation and protonation states of C P . Helix-coil transitionsusually involve overcoming high energy barriers, thus conventional molecular dynamics (MD)simulations become insufficient to study this kind of conformational rearrangements. Tosurpass this obstacle, we employ the accelerated molecular dynamics scheme, an enhancedsampling technique that extends the effective simulation time scale to long microseconds.Preliminary results suggest that the FF➝LU transition has a higher associated energybarrier than the refolding LU➝FF process in agreement with the experimental low catalyticrate constant of Ec TPx. Furthermore, in silico designed point-mutants of the αH3 helixenhanced locally unfolding events, suggesting that the native interactions in the active site ofthe FF native substate are not optimized for fully speed-up the conformational transition.AcknowledgementsThis work was supported by grants from the Universidad de Buenos Aires N°20020130100468BA, theAgencia Nacional de Promoción Científica y Tecnológica PICT N°2013-0983 and PICT-CABBION°2013-3362 and the Consejo Nacional de Investigaciones Científicas y Técnicas PIP 2010-2015