title: Chemical denaturation of a homodimeric lysine-49 phospholipase A2: a stable dimer interface and a native monomeric intermediate.
abstract: Bothropstoxin I (4BthTx-I) is a homodimeric lysine-49 (Lys49) phospholipase A(2) isolated from Bothrops jararacussu venom, which damages liposome membranes via a Ca(2+)-independent mechanism. The stability of the BthTx-I homodimer was evaluated by equilibrium chemical denaturation with guanidinium hydrochloride monitored by changes in the intrinsic tryptophan fluorescence anisotropy, far-UV circular dichroism, dynamic light scattering, and 1-anilinonaphthalene-8-sulfonate binding. Unfolding of the BthTx-I dimer proceeds via a monomeric intermediate with native-like structure, with Gibbs free energy (DeltaG(0)) values of 10.0 and 7.2 kcal mol(-1) for the native dimer-to-native monomer and native-to-denatured monomer transitions, respectively. The experimentally determined DeltaG(0) value for the dimer-to-native monomer transition is higher than the value expected for an interaction dominated by hydrophobic forces, and suggests that an unusually high propensity of hydrogen-bonded side chains found at the BthTx-I homodimer interface make a significant contribution to dimer stability.
authors.affiliation: Departamento de Bioqui;mica e Immunologia, FMRP-USP, Universidade de São Paulo, Ribeirão Preto-SP, Brazil.
authors.forename: Roberto
authors.initials: R
authors.lastname: Ruller
authors.forename: Tatiana Lopes
authors.initials: TL
authors.lastname: Ferreira
authors.forename: Arthur H C
authors.initials: AH
authors.lastname: de Oliveira
authors.forename: Richard J
authors.initials: RJ
authors.lastname: Ward
mesh_terms: D000818:Animals; D017837:Bothrops; D002942:Circular Dichroism; D003435:Crotalid Venoms / Q000737:chemistry*; D019281:Dimerization; D004795:Enzyme Stability; D005454:Fluorescence Polarization; D005456:Fluorescent Dyes; D008027:Light; D008239:Lysine; D010741:Phospholipases A / Q000737:chemistry* / Q000302:isolation & purification; D054467:Phospholipases A2; D011487:Protein Conformation; D011489:Protein Denaturation; D012542:Scattering, Radiation
chemical_list: D003435:Crotalid Venoms; D005456:Fluorescent Dyes; D010741:Phospholipases A; D054467:Phospholipases A2; D008239:Lysine