Chloroplast 2-Cys Peroxiredoxin. Modulation of chaperone activity by Dithiotreitol and ATP-Mg2+

MARTÍN ARÁN, PATRICIO CRAIG, DANIEL CAPORALETTI, ALEJANDRO SENN AND RICARDO A. WOLOSIUK.

Iguazú, Argentina

Congreso; XL SAIB ANNUAL MEETING; 2004

Sociedad Argentina de Investigación en Bioquímica y Biología Molecular

Peroxiredoxins (Prxs) are peroxidases linked to diverse cellular functions that reduce H2O2, peroxinitrite, and organic hydroperoxide. These ubiquitous antioxidant proteins contain one and two conserved cysteine residues, 1-Cys and 2-Cys Prx, respectively, that are functional in the peroxidatic activity using reduced thioredoxin as electron donor. X-ray crystal structures of human erythrocytes and Crithidia fasciculata 2-Cys Prx evinced the noncovalent association of five homodimers whose constituent monomers are linked via a disulfide bond; i.e. [(Prx)2]5. To estimate the chaperone activity of rapeseed chloroplast 2- Cys Prx, we followed the denaturation of citrate synthase and glyceraldehyde 3-P dehydrogenase by light scattering. 2-Cys Prx prevented the thermal aggregation of both enzymes in a process that was enhanced by reduction with dithiothreitol (DTT). Size exclusion cromatography and light scattering studies showed that chloroplast 2-Cys Prx adopted a decameric structure when DTT cleaved the disulfide bond of the dimer. Moreover, both ATP and Mg2+ inhibited the chaperone activity of chloroplast 2-Cys Prx. Apparently, the concerted action of ATP and Mg2+ caused the precipitation of chloroplast 2-Cys Prx and, in so doing, impaired the protective capacity. These results indicate that, in vitro, chloroplast 2-Cys Prx is a chaperone whose activity could be modulated via changes in the quaternary structure by ATP and the reduction of the intrinsic disulfide bond.