title: Evaluation of the conformational equilibrium of reduced hen egg lysozyme by antibodies to the native form.
abstract: To evaluate the conformation of reduced HEL, the monoclonal antibodies HyC1 and HyC2, which recognize different conformational epitopes on native hen egg lysozyme (HEL), were used, and the kinetics of their interactions with native HEL, S-1,2-dicarboxyethylated HEL (DCE-HEL), and carboxymethylated Cys6 and Cys127 HEL (CM(6,127)-HEL) were assessed using surface plasmon resonance. Although their association rate constants differed 10(5)-fold, their dissociation rate constants were essentially the same, suggesting that DCE-HEL and CM(6,127)-HEL possess conformations similar to that of native HEL when they bind antibodies. We considered that the ratio of the association rate constant of reduced HEL to native HEL represents the proportion of the native format determinant in equilibrium. Reduction of the Cys6-Cys127 disulfide bond would transform the epitope recognized by HyC1 into a non-native conformation similar to that of DCE-HEL. We show that monoclonal antibodies provide a sensitive tool for evaluation of the structural and hydrodynamic changes of proteins.
authors: Graduate School of Life and Environmental Sciences, Kyoto Prefectural University, 1-5 Hangi-cho, Shimogamo, Sakyo-ku, Kyoto 606-8522, Japan.
authors: Masayuki
authors: M
authors: Oda
authors: Aki
authors: A
authors: Kitai
authors: Akikazu
authors: A
authors: Murakami
authors: Miyuki
authors: M
authors: Nishimura
authors: Takatoshi
authors: T
authors: Ohkuri
authors: Yoshito
authors: Y
authors: Abe
authors: Tadashi
authors: T
authors: Ueda
authors: Haruki
authors: H
authors: Nakamura
authors: Takachika
authors: T
authors: Azuma
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000939:Epitopes / Q000276:immunology*; D007700:Kinetics; D009113:Muramidase / Q000737:chemistry* / Q000276:immunology* / Q000378:metabolism; D010084:Oxidation-Reduction; D011487:Protein Conformation; D020673:Protein Renaturation; D012996:Solutions; D020349:Surface Plasmon Resonance
chemical_list: D000911:Antibodies, Monoclonal; D000939:Epitopes; D012996:Solutions; C421864:hen egg lysozyme; D009113:Muramidase