title: A novel method of preparing the monoform structure of catalytic antibody light chain.
abstract: Along with the development of antibody drugs and catalytic antibodies, the structural diversity (heterogeneity) of antibodies has been given attention. For >20 yr, detailed studies on the subject have not been conducted, because the phenomenon presents many difficult and complex problems. Structural diversity provides some (or many) isoforms of an antibody distinguished by different charges, different molecular sizes, and modifications of amino acid residues. For practical use, the antibody and the subunits must have a defined structure. In recent work, we have found that the copper (Cu) ion plays a substantial role in solving the diversity problem. In the current study, we used several catalytic antibody light chains to examine the effect of the Cu ion. In all cases, the different electrical charges of the molecule converged to a single charge, giving 1 peak in cation-exchange chromatography, as well as a single spot in 2-dimensional gel electrophoresis. The Cu-binding site was investigated by using mutagenesis, ultraviolet-visible spectroscopy, atomic force microscope analysis, and molecular modeling, which suggested that histidine and cysteine residues close to the C-terminus are involved with the binding site. The constant region domain of the antibody light chain played an important role in the heterogeneity of the light chain. Our findings may be a significant tool for preparing a single defined, not multiple, isoform structure.
authors.affiliation: *Research Promotion Institute and Department of Applied Chemistry, Faculty of Engineering, Oita University, Oita, Japan; Nanotechnology Laboratory, Institute of Systems, Information Technologies, and Nanotechnologies (ISIT), Fukuoka, Japan; Graduate School of System Life Science, Kyushu University, Fukuoka, Japan; Tottori College of Nursing, Tottori, Japan; and High Energy Accelerator Research Organization, Tsukuba, Japan e-hifumi@oita-u.ac.jp.
authors.forename: Emi
authors.initials: E
authors.lastname: Hifumi
authors.affiliation: *Research Promotion Institute and Department of Applied Chemistry, Faculty of Engineering, Oita University, Oita, Japan; Nanotechnology Laboratory, Institute of Systems, Information Technologies, and Nanotechnologies (ISIT), Fukuoka, Japan; Graduate School of System Life Science, Kyushu University, Fukuoka, Japan; Tottori College of Nursing, Tottori, Japan; and High Energy Accelerator Research Organization, Tsukuba, Japan.
authors.forename: Shingo
authors.initials: S
authors.lastname: Matsumoto
authors.affiliation: *Research Promotion Institute and Department of Applied Chemistry, Faculty of Engineering, Oita University, Oita, Japan; Nanotechnology Laboratory, Institute of Systems, Information Technologies, and Nanotechnologies (ISIT), Fukuoka, Japan; Graduate School of System Life Science, Kyushu University, Fukuoka, Japan; Tottori College of Nursing, Tottori, Japan; and High Energy Accelerator Research Organization, Tsukuba, Japan.
authors.forename: Hiroki
authors.initials: H
authors.lastname: Nakashima
authors.affiliation: *Research Promotion Institute and Department of Applied Chemistry, Faculty of Engineering, Oita University, Oita, Japan; Nanotechnology Laboratory, Institute of Systems, Information Technologies, and Nanotechnologies (ISIT), Fukuoka, Japan; Graduate School of System Life Science, Kyushu University, Fukuoka, Japan; Tottori College of Nursing, Tottori, Japan; and High Energy Accelerator Research Organization, Tsukuba, Japan.
authors.forename: Shogo
authors.initials: S
authors.lastname: Itonaga
authors.affiliation: *Research Promotion Institute and Department of Applied Chemistry, Faculty of Engineering, Oita University, Oita, Japan; Nanotechnology Laboratory, Institute of Systems, Information Technologies, and Nanotechnologies (ISIT), Fukuoka, Japan; Graduate School of System Life Science, Kyushu University, Fukuoka, Japan; Tottori College of Nursing, Tottori, Japan; and High Energy Accelerator Research Organization, Tsukuba, Japan.
authors.forename: Mitsue
authors.initials: M
authors.lastname: Arakawa
authors.affiliation: *Research Promotion Institute and Department of Applied Chemistry, Faculty of Engineering, Oita University, Oita, Japan; Nanotechnology Laboratory, Institute of Systems, Information Technologies, and Nanotechnologies (ISIT), Fukuoka, Japan; Graduate School of System Life Science, Kyushu University, Fukuoka, Japan; Tottori College of Nursing, Tottori, Japan; and High Energy Accelerator Research Organization, Tsukuba, Japan.
authors.forename: Yoshiki
authors.initials: Y
authors.lastname: Katayama
authors.affiliation: *Research Promotion Institute and Department of Applied Chemistry, Faculty of Engineering, Oita University, Oita, Japan; Nanotechnology Laboratory, Institute of Systems, Information Technologies, and Nanotechnologies (ISIT), Fukuoka, Japan; Graduate School of System Life Science, Kyushu University, Fukuoka, Japan; Tottori College of Nursing, Tottori, Japan; and High Energy Accelerator Research Organization, Tsukuba, Japan.
authors.forename: Ryuichi
authors.initials: R
authors.lastname: Kato
authors.affiliation: *Research Promotion Institute and Department of Applied Chemistry, Faculty of Engineering, Oita University, Oita, Japan; Nanotechnology Laboratory, Institute of Systems, Information Technologies, and Nanotechnologies (ISIT), Fukuoka, Japan; Graduate School of System Life Science, Kyushu University, Fukuoka, Japan; Tottori College of Nursing, Tottori, Japan; and High Energy Accelerator Research Organization, Tsukuba, Japan.
authors.forename: Taizo
authors.initials: T
authors.lastname: Uda
mesh_terms: D017151:Antibodies, Catalytic / Q000737:chemistry* / Q000302:isolation & purification; D001666:Binding Sites, Antibody; D003300:Copper / Q000737:chemistry*; D006801:Humans; D007147:Immunoglobulin Light Chains / Q000737:chemistry* / Q000302:isolation & purification
chemical_list: D017151:Antibodies, Catalytic; D007147:Immunoglobulin Light Chains; D003300:Copper
keywords: 2D electrophoresis; charge heterogeneity; copper ion; pI