title: Role of the mobility of antigen binding site in high affinity antibody elucidated by surface plasmon resonance.
abstract: Antibody is known to exhibit conformational change in the antigen recognition site after forming the initial complex. This structural change, which is widely known as "induced fit", is believed to be critical for high affinity (Kdof nM range) of antigen-antibody interaction. Elucidation of this 'induced fit' process is essential for rational design of high affinity antibody, while it is prevented by limitation of the available biophysical and biochemical data of the initial complex. Here, we performed kinetic and thermodynamic analysis of the interaction between single-chain variable fragment (denoted as scFv) of 64M5 antibody and a (6-4) photoproduct by using surface plasmon resonance (denoted as SPR). It revealed that the 64M5scFv associates the (6-4) photoproduct at initial step by hydrophobic interactions, and enthalpy-driving interactions, hydrogen bonds and van der Waals interactions, were formed by second step structural rearrangement. Furthermore, mutational analysis revealed that the mobility of the antigen-binding site is critical for the second step. It could be assumed that optimization of the mobility of the antigen recognition site is a clue for rational design of high affinity antibody.
authors: Department of Analytical and Biophysical Chemistry, Graduate School of Pharmaceutical Sciences, Kumamoto University, 5-1 Oe-honmachi, Chuo-ku, Kumamoto 862-0973, Japan.
authors: Natsuki
authors: N
authors: Fukuda
authors: Department of Analytical and Biophysical Chemistry, Graduate School of Pharmaceutical Sciences, Kumamoto University, 5-1 Oe-honmachi, Chuo-ku, Kumamoto 862-0973, Japan.
authors: Yoshiaki
authors: Y
authors: Suwa
authors: Department of Analytical and Biophysical Chemistry, Graduate School of Pharmaceutical Sciences, Kumamoto University, 5-1 Oe-honmachi, Chuo-ku, Kumamoto 862-0973, Japan.
authors: Makiyo
authors: M
authors: Uchida
authors: Department of Analytical and Biophysical Chemistry, Graduate School of Pharmaceutical Sciences, Kumamoto University, 5-1 Oe-honmachi, Chuo-ku, Kumamoto 862-0973, Japan.
authors: Yoshihiro
authors: Y
authors: Kobashigawa
authors: Labratory of Physical Biochemistry, Faculty of Pharmaceutical Sciences, Tokyo University of Science, 2641 Yamazaki, Noda, Chiba 278-8510, Japan.
authors: Hideshi
authors: H
authors: Yokoyama
authors: Department of Analytical and Biophysical Chemistry, Graduate School of Pharmaceutical Sciences, Kumamoto University, 5-1 Oe-honmachi, Chuo-ku, Kumamoto 862-0973, Japan morioka@kumamoto-u.ac.jp.
authors: Hiroshi
authors: H
authors: Morioka
mesh_terms: D000818:Animals; D001666:Binding Sites, Antibody; D051379:Mice; D057127:Single-Chain Antibodies / Q000737:chemistry*; D020349:Surface Plasmon Resonance
chemical_list: D057127:Single-Chain Antibodies
keywords: (6-4) photoproduct; scFv; structural rearrangement; surface plasmon resonance; thermodynamics parameter