Heat-induced Irreversible Denaturation of the Camelid Single Domain VHH Antibody Is Governed by Chemical Modifications

Akazawa, Yoko; Takashima, Mizuki; Lee, Young-Ho; Ikegami, Takahisa; Goto, Yuji; Uegaki, Koichi; Hagihara, Yoshihisa

doi:https://doi.org/10.1074/jbc.M113.534222

Heat-induced Irreversible Denaturation of the Camelid Single Domain VHH Antibody Is Governed by Chemical Modifications

Akazawa, Yoko; Takashima, Mizuki; Lee, Young-Ho et al.

Journal of Biological Chemistry, 2014, 289(22), 15666-15679

アクセス数:324件（2025-05-13 06:35 集計）

固定URL: https://hdl.handle.net/11094/71278

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タイトル	Heat-induced Irreversible Denaturation of the Camelid Single Domain VHH Antibody Is Governed by Chemical Modifications
著者	NRID 1000050549897 1000050549897 著者名 Akazawa, Yoko 著者の別表記 Ogawa, Yoko
	著者名 Takashima, Mizuki
	NRID 1000070589431 1000070589431 著者名 Lee, Young-Ho
	NRID 1000020283939 1000020283939 著者名 Ikegami, Takahisa
	NRID 1000040153770 1000040153770 researchmap permalink read0014440 read0014440 著者名 Goto, Yuji
	NRID 1000000356544 1000000356544 著者名 Uegaki, Koichi
	NRID 1000050357761 1000050357761 著者名 Hagihara, Yoshihisa
内容	This research was originally published in the Journal of Biological Chemistry. Yoko Akazawa-Ogawa, Mizuki Takashima, Young-Ho Lee, Takahisa Ikegami, Yuji Goto, Koichi Uegaki and Yoshihisa Hagihara. Heat-induced Irreversible Denaturation of the Camelid Single Domain VHH Antibody Is Governed by Chemical Modifications. J. Biol. Chem. 2014; 289, 15666–15679. © the American Society for Biochemistry and Molecular Biology
抄録	The variable domain of camelid heavy chain antibody (VHH) is highly heat-resistant and is therefore ideal for many applications. Although understanding the process of heat-induced irreversible denaturation is essential to improve the efficacy of VHH, its inactivation mechanism remains unclear. Here, we showed that chemical modifications predominantly governed the irreversible denaturation of VHH at high temperatures. After heat treatment, the activity of VHH was dependent only on the incubation time at 90 °C and was insensitive to the number of heating (90 °C)-cooling (20 °C) cycles, indicating a negligible role for folding/unfolding intermediates on permanent denaturation. The residual activity was independent of concentration; therefore, VHH lost its activity in a unimolecular manner, not by aggregation. A VHH mutant lacking Asn, which is susceptible to chemical modifications, had significantly higher heat resistance than did the wild-type protein, indicating the importance of chemical modifications to VHH denaturation.
出版者	American Society for Biochemistry and Molecular Biology
収録物名	Journal of Biological Chemistry
巻(号)	289(22)
ページ	15666-15679
刊行年月	2014-05
言語	英語
ハンドルURL	https://hdl.handle.net/11094/71278
eISSN	1083351X
NCID	AA1202447X
関連情報 (isIdenticalTo)	DOI (出版社版) https://doi.org/10.1074/jbc.M113.534222
アクセス権	open access
出版タイプ	Version of Record
カテゴリ	学術雑誌論文 Journal Article

資源タイプ	学術雑誌論文
ローカル資源タイプ	学術雑誌論文
DCMI資源タイプ	text
DCTERMS.bibliographicCitation	Journal of Biological Chemistry. 2014, 289(22), p. 15666-15679
DC.title	Heat-induced Irreversible Denaturation of the Camelid Single Domain VHH Antibody Is Governed by Chemical Modifications
DC.creator	Akazawa, Yoko
	Takashima, Mizuki
	Lee, Young-Ho
	Ikegami, Takahisa
	Goto, Yuji
	Uegaki, Koichi
	Hagihara, Yoshihisa
DC.creator	Ogawa, Yoko
DC.publisher	American Society for Biochemistry and Molecular Biology
DC.language' scheme='DCTERMS.RFC1766	英語
DC.type' scheme='DCTERMS.DCMIType	text
DCTERMS.issued' scheme='DCTERMS.W3CDTF	2014-05
DC.identifier	https://doi.org/10.1074/jbc.M113.534222
DC.description	This research was originally published in the Journal of Biological Chemistry. Yoko Akazawa-Ogawa, Mizuki Takashima, Young-Ho Lee, Takahisa Ikegami, Yuji Goto, Koichi Uegaki and Yoshihisa Hagihara. Heat-induced Irreversible Denaturation of the Camelid Single Domain VHH Antibody Is Governed by Chemical Modifications. J. Biol. Chem. 2014; 289, 15666–15679. © the American Society for Biochemistry and Molecular Biology
DCTERMS.abstract	The variable domain of camelid heavy chain antibody (VHH) is highly heat-resistant and is therefore ideal for many applications. Although understanding the process of heat-induced irreversible denaturation is essential to improve the efficacy of VHH, its inactivation mechanism remains unclear. Here, we showed that chemical modifications predominantly governed the irreversible denaturation of VHH at high temperatures. After heat treatment, the activity of VHH was dependent only on the incubation time at 90 °C and was insensitive to the number of heating (90 °C)-cooling (20 °C) cycles, indicating a negligible role for folding/unfolding intermediates on permanent denaturation. The residual activity was independent of concentration; therefore, VHH lost its activity in a unimolecular manner, not by aggregation. A VHH mutant lacking Asn, which is susceptible to chemical modifications, had significantly higher heat resistance than did the wild-type protein, indicating the importance of chemical modifications to VHH denaturation.
DC.format	71278
citation_title	Heat-induced Irreversible Denaturation of the Camelid Single Domain VHH Antibody Is Governed by Chemical Modifications
citation_author	Akazawa, Yoko
	Takashima, Mizuki
	Lee, Young-Ho
	Ikegami, Takahisa
	Goto, Yuji
	Uegaki, Koichi
	Hagihara, Yoshihisa
citation_publisher	American Society for Biochemistry and Molecular Biology
citation_language	英語
citation_date	2014-05
citation_journal_title	Journal of Biological Chemistry
citation_volume	289
citation_issue	22
citation_firstpage	15666
citation_lastpage	15679
citation_doi	https://doi.org/10.1074/jbc.M113.534222
citation_public_url	https://hdl.handle.net/11094/71278

Heat-induced Irreversible Denaturation of the Camelid Single Domain VHH Antibody Is Governed by Chemical Modifications

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