title: Acid- and pressure-induced (un)folding of yeast glutathione reductase: competition between protein oligomerization and aggregation.
abstract: Glutathione reductase (GR) is a homodimeric flavoenzyme involved in cellular defense against oxidative stress. In the present study, we have used a combination of acidic pH and hydrostatic pressure to investigate the (un)folding transition of yeast GR. Our results indicate that at pH 2 a distinct partially folded state is stabilized, as judged by intrinsic fluorescence, bis ANS binding and circular dichroism (CD) analysis. Further characterization of this partially folded state by size exclusion chromatography revealed that it corresponds to expanded GR monomers. CD analysis at pH 2 showed a significant loss of secondary structure. The partially folded GR monomers stabilized at pH 2 were fully and reversibly unfolded using hydrostatic pressure (up to 3.5kbar) as a thermodynamic perturbant. By contrast, return to physiological pH after exposure to acidic pH led to a competing reaction between refolding dimerization and aggregation of GR. These results support the notion that a partially folded intermediate state is not only critical for folding of GR but also appears to be a seed for protein aggregation.
authors.affiliation: Instituto de Bioquímica Médica, Programa de Bioquímica e Biofísica Celular, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil.
authors.forename: Ana Cristina S
authors.initials: AC
authors.lastname: Morais
authors.forename: Alex
authors.initials: A
authors.lastname: Chapeaurouge
authors.forename: Sérgio T
authors.initials: ST
authors.lastname: Ferreira
mesh_terms: D000143:Acids / Q000494:pharmacology*; D000817:Anilino Naphthalenesulfonates; D001274:Atmospheric Pressure; D019281:Dimerization; D005980:Glutathione Reductase / Q000737:chemistry* / Q000378:metabolism*; D011487:Protein Conformation; D017510:Protein Folding; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D013050:Spectrometry, Fluorescence
chemical_list: D000143:Acids; D000817:Anilino Naphthalenesulfonates; C035763:5,5'-bis(8-(phenylamino)-1-naphthalenesulfonate); D005980:Glutathione Reductase