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

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Identifier to cite or link to this item: https://hdl.handle.net/11094/71278

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Title
Heat-induced Irreversible Denaturation of the Camelid Single Domain VHH Antibody Is Governed by Chemical Modifications
Creator
NRID
1000050549897 1000050549897
CreatorName
Akazawa, Yoko
Creator.Alternative
Ogawa, Yoko
CreatorName
Takashima, Mizuki
NRID
1000070589431 1000070589431
CreatorName
Lee, Young-Ho
NRID
1000020283939 1000020283939
CreatorName
Ikegami, Takahisa
NRID
1000040153770 1000040153770
researchmap permalink
read0014440 read0014440
CreatorName
Goto, Yuji
NRID
1000000356544 1000000356544
CreatorName
Uegaki, Koichi
NRID
1000050357761 1000050357761
CreatorName
Hagihara, Yoshihisa
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
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.
Publisher
American Society for Biochemistry and Molecular Biology
Source Title
Journal of Biological Chemistry
Volume (Issue)
289(22)
Page
15666-15679
Date of Issued
2014-05
Language
English
Handle URL
https://hdl.handle.net/11094/71278
eISSN
1083351X
NCID
AA1202447X
Relation.isIdenticalTo
DOI (Publisher Version)
https://doi.org/10.1074/jbc.M113.534222
Access Rights
open access
oaire:version
Version of Record
Category
学術雑誌論文 Journal Article