title: Structural characterization of acid-induced intermediates of human glutathione transferase P1-1.
abstract: The acid denaturation of human glutathione transferase P1-1 (hGSTP1-1) has been performed to investigate the unfolding intermediates of the protein and their possible involvement in the refolding mechanism. The acid-induced structures of GSTP1-1 have been characterized by activity, gel filtration, intrinsic fluorescence and far-u.v. circular dichroism (CD) techniques. Because of the non-identity of the different transitions monitored, the acid denaturation of hGSTP1-1 appears to be a multistep process during which several intermediates coexist in equilibrium. The dependence of inactivation on the protein concentration, as well as gel-filtration experiments, indicate that the inactivation transition, centred at about pH 4.0, corresponds to the monomerization of the protein. At pH 2.0, when the enzyme is completely inactive, the protein retains a small, but significant, amount of secondary structure. This means that the dimeric arrangement of the molecule is important for maintaining the native-like secondary structure of the monomer. The results show that, at low pH, the compact state of the GST monomer, even upon the addition of salts, does not possess native-like secondary structure as described for many monomeric proteins (molten globule). In the presence of physiological concentrations of salts, the protein solution at pH 2.0 leads to a dead-end aggregation process, suggesting that this compact state cannot represent a productive intermediate of the refolding pathway.
authors.affiliation: Dipartimento di Scienze Biomediche, Università G. D'Annunzio, Via dei Vestini, 31, 66100 Chieti, Italy.
authors.forename: B
authors.initials: B
authors.lastname: Dragani
authors.forename: R
authors.initials: R
authors.lastname: Cocco
authors.forename: D R
authors.initials: DR
authors.lastname: Principe
authors.forename: M
authors.initials: M
authors.lastname: Cicconetti
authors.forename: A
authors.initials: A
authors.lastname: Aceto
mesh_terms: D002942:Circular Dichroism; D019343:Citric Acid / Q000494:pharmacology; D051549:Glutathione S-Transferase pi / Q000737:chemistry* / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D011487:Protein Conformation; D011489:Protein Denaturation / Q000187:drug effects; D017510:Protein Folding / Q000187:drug effects
chemical_list: D019343:Citric Acid; D051549:Glutathione S-Transferase pi