Accessing to Local Unfolding Events that take place during the Catalytic Cycle of E. coli Thiolperoxidase

VAZQUEZ DS; AGUDELO WA; FERRER-SUETA G; SANTOS J

oz do Iguaçu

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

Brazilian Society for Biochemistry and Molecular Biology

Introduction and objectives: Peroxiredoxins (in particular the TPX subfamily) are a ubiquitous family of enzymes which reduce different kind of peroxides via a reactive cysteine. Prxs require a key peroxidatic cysteine (CP) that, in an active fully folded (FF) conformation, is oxidized to sulfenic acid (CysOH) and then, after a critically local unfolding step (LU), forms a disulfide bond with a second resolving cysteine (CR) (Fig.1A). In TPX the FF→LU transition involves the helix α2 and α3 containing the CP and CR cysteines respectively (Fig.1B and 1C). We hypothesize that the α3 helix has an intrinsic tenency to unfold and the energetic barrier involved in this process limits the catalityc ratio of the cycle. In order to study the preceding events and the structural determinants of the CP-CR disulfide bond linkage, we performed a biased conformational searching profiles followed by PCA analysis and the experimental measurement of the stability of each conformation. Materials and methods: Common and accelerated MD simulations were carried out and analyzed in the AMBER14-GPU package using TIP3P waters at 300 K and NVT ensemble. PCA analysis were performed to identifiy and clusterize different dynamic-based populations. 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, due to the higher α-value needed to reach to the other basin.