title: The effect of high pH upon diphtheria toxin conformation and model membrane association: role of partial unfolding.
abstract: Penetration of membranes by diphtheria toxin in vivo is at least partially triggered by a low pH-induced conformational change occurring within the lumen of an acidic organelle. In order to gain insight into the nature of this change the behavior of the toxin at high pH was characterized and compared to that previously determined at low pH. We find that near pH 10.5 a major conformational change occurs. This change is accompanied by a marked decrease in fluorescence intensity, a red shift in fluorescence emission maximum, and increased susceptibility of protein fluorescence to acrylamide quenching. Differential scanning calorimetry shows that the high pH conformational change involves a cooperative endothermic unfolding transition. These changes at high pH are very similar to those induced by low pH, supporting the conclusion that the changes at low pH also involve a denaturation-like process. In addition, at high pH the toxin gains the ability to bind to model membranes, again similar to its behavior at low pH. On the basis of these studies we conclude that exposure of hydrophobic sequences due to partial unfolding is one dominating component in inducing hydrophobic behavior at both high and low pH, but that at low pH Asp/Glu protonation also contributes to hydrophobicity.
authors.affiliation: Department of Biochemistry and Cell Biology, S.U.N.Y., Stony Brook 11794-5215.
authors.forename: J
authors.initials: J
authors.lastname: Kieleczawa
authors.forename: J M
authors.initials: JM
authors.lastname: Zhao
authors.forename: C L
authors.initials: CL
authors.lastname: Luongo
authors.forename: L Y
authors.initials: LY
authors.lastname: Dong
authors.forename: E
authors.initials: E
authors.lastname: London
mesh_terms: D002152:Calorimetry, Differential Scanning; D004167:Diphtheria Toxin / Q000737:chemistry*; D006863:Hydrogen-Ion Concentration; D008567:Membranes, Artificial; D008823:Micelles; D010539:Permeability; D011487:Protein Conformation; D011489:Protein Denaturation; D013050:Spectrometry, Fluorescence
chemical_list: D004167:Diphtheria Toxin; D008567:Membranes, Artificial; D008823:Micelles