Fc domain mediated self-association of an IgG1 monoclonal antibody under a low ionic strength condition(ENZYMOLOGY, PROTEIN ENGINEERING, AND ENZYME TECHNOLOGY)
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概要
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Recently, we reported that IgG1 monoclonal antibody A (MAb A) underwent liquid-liquid phase separation and separated into light and heavy phases under a low ionic strength condition. The liquid-liquid phase separation was induced due to self-association of MAb A in the heavy phase when the initial concentration of MAb A was between the two critical concentrations [Nishi et al., Pharm. Res., 27, 1348-1360 (2010)]. Here, we determined the interaction site of MAb A by using proteolytic Fab and Fc fragments of MAb A. The mean hydrodynamic diameter of the Fc fragment increased in a low ionic strength buffer, and furthermore the SPR measurement detected interactions of the Fc fragment with both whole MAb A and the Fc fragment, whereas the Fab fragment interacted with neither whole MAb A nor the Fc fragment. No binding was detected under an isotonic ionic strength condition. Zeta potential of MAb A was significant positive below pH5.5 and negative above pH6.5. Between pH5.5 and 6.5 where the phase separation is significantly induced, MAb A had only a small positive or negative net charge. The isothermal titration calorimetry dilution method revealed that dissociation of MAb A accompanied endothermic heat changes, suggesting that intermolecular interactions among MAb A molecules were attributed to the enthalpically driven process. These results suggest that liquid-liquid phase separation of MAb A is mediated by a weak electrostatic intermolecular interaction among MAb A molecules mainly at Fc portions.
著者
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Fukui Kiichi
Department Of Biotechnology Graduate School Of Engineering Osaka University
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Fukui Kiichi
Department Of Biotechnology Faculty Of Engineering Graduate School Of Osaka University
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Uchiyama Susumu
Department Of Biotechnology Graduate School Of Engineering Osaka University
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Wakiyama Naoki
Formulation Technology Research Laboratories, Daiichi-Sankyo Co., Ltd.
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Nishi Hirotaka
Formulation Technology Research Laboratories, Daiichi Sankyo Co., Ltd.
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Miyajima Makoto
Formulation Technology Research Laboratories, Daiichi Sankyo Co., Ltd.
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Kubota Kei
Analytical & Quality Evaluation Research Laboratories, Daiichi Sankyo Co., Ltd.
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Hasegawa Jun
Analytical & Quality Evaluation Research Laboratories, Daiichi Sankyo Co., Ltd.
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Wakiyama Naoki
Formulation Technology Research Laboratories Daiichi Sankyo Co. Ltd.
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Miyajima Makoto
Formulation Technology Research Laboratories Daiichi Sankyo Co. Ltd.
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Kubota Kei
Analytical & Quality Evaluation Research Laboratories Daiichi Sankyo Co. Ltd.
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Nishi Hirotaka
Formulation Technology Research Laboratories Daiichi Sankyo Co. Ltd.:department Of Biotechnology Gra
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Hasegawa Jun
Analytical & Quality Evaluation Research Laboratories Daiichi Sankyo Co. Ltd.
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