Functional Divergence in the Affinity and Stability of Non-Canonical Cysteines and Non-Canonical Disulfide Bonds: Insights from a VHH and VNAR Study

doi:10.3390/ijms25189801

IOCAS-IR > 实验海洋生物学重点实验室

	Functional Divergence in the Affinity and Stability of Non-Canonical Cysteines and Non-Canonical Disulfide Bonds: Insights from a VHH and VNAR Study
	Xu, Mingce 1,2,3; Zhao, Zheng 4,5; Deng, Penghui 1,2,3; Sun, Mengsi 6; Chiu, Cookson K. C.6; Wu, Yujie 6; Wang, Hao 1,2,7; Bi, Yunchen 1,2,3,7
	2024-09-01
发表期刊	INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES
ISSN	1661-6596
卷号	25 期号:18 页码:16
通讯作者	Wang, Hao([email protected]) ; Bi, Yunchen([email protected])
摘要	Single-domain antibodies, including variable domains of the heavy chains of heavy chain-only antibodies (VHHs) from camelids and variable domains of immunoglobulin new antigen receptors (VNARs) from cartilaginous fish, show the therapeutic potential of targeting antigens in a cytosol reducing environment. A large proportion of single-domain antibodies contain non-canonical cysteines and corresponding non-canonical disulfide bonds situated on the protein surface, rendering them vulnerable to environmental factors. Research on non-canonical disulfide bonds has been limited, with a focus solely on VHHs and utilizing only cysteine mutations rather than the reducing agent treatment. In this study, we examined an anti-lysozyme VNAR and an anti-BC2-tag VHH, including their non-canonical disulfide bond reduced counterparts and non-canonical cysteine mutants. Both the affinity and stability of the VNARs and VHHs decreased in the non-canonical cysteine mutants, whereas the reduced-state samples exhibited decreased thermal stability, with their affinity remaining almost unchanged regardless of the presence of reducing agents. Molecular dynamics simulations suggested that the decrease in affinity of the mutants resulted from increased flexibility of the CDRs, the disappearance of non-canonical cysteine-antigen interactions, and the perturbation of other antigen-interacting residues caused by mutations. These findings highlight the significance of non-canonical cysteines for the affinity of single-domain antibodies and demonstrate that the mutation of non-canonical cysteines is not equivalent to the disruption of non-canonical disulfide bonds with a reducing agent when assessing the function of non-canonical disulfide bonds.
关键词	single-domain antibodies disulfide bonds cysteines nanobodies intrabodies
DOI	10.3390/ijms25189801
收录类别	SCI
语种	英语
资助项目	National Natural Science Foundation of China; Taishan Scholar Program of Shandong Province[tstp20240840]; Users with Excellence Program of Hefei Science Center, CAS[2020HSC-UE018]; Research on Simulation Technology and Device of Key Processes of Typical Marine Ecological Disasters in the Pre-Research Project of Major Scientific Facilities in Shandong Province[DKXZZ202205]; [42376136]; [42306144]
WOS研究方向	Biochemistry & Molecular Biology ; Chemistry
WOS类目	Biochemistry & Molecular Biology ; Chemistry, Multidisciplinary
WOS记录号	WOS:001323829800001
出版者	MDPI
WOS关键词	DOMAIN ANTIBODIES ; NANOBODIES ; SELECTION ; DYNAMICS
引用统计	被引频次：1[WOS] [WOS记录] [WOS相关记录]
文献类型	期刊论文
条目标识符	http://ir.qdio.ac.cn/handle/337002/199244
专题	实验海洋生物学重点实验室
通讯作者	Wang, Hao; Bi, Yunchen
作者单位	1.Chinese Acad Sci, Qingdao 266071, Peoples R China 2.Chinese Acad Sci, Inst Oceanol, Ctr Ocean Mega Sci, Shandong Prov Key Lab Expt Marine Biol, Qingdao 266071, Peoples R China 3.Univ Chinese Acad Sci, Beijing 100049, Peoples R China 4.Univ Virginia, Sch Data Sci, Charlottesville, VA 22904 USA 5.Univ Virginia, Dept Biomed Engn, Charlottesville, VA 22904 USA 6.Shenzhen Bay Lab, Shenzhen 518055, Peoples R China 7.Qingdao Marine Sci & Technol Ctr, Lab Marine Biol & Biotechnol, Qingdao 266071, Peoples R China
推荐引用方式 GB/T 7714	Xu, Mingce,Zhao, Zheng,Deng, Penghui,et al. Functional Divergence in the Affinity and Stability of Non-Canonical Cysteines and Non-Canonical Disulfide Bonds: Insights from a VHH and VNAR Study[J]. INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES,2024,25(18):16.
APA	Xu, Mingce.,Zhao, Zheng.,Deng, Penghui.,Sun, Mengsi.,Chiu, Cookson K. C..,...&Bi, Yunchen.(2024).Functional Divergence in the Affinity and Stability of Non-Canonical Cysteines and Non-Canonical Disulfide Bonds: Insights from a VHH and VNAR Study.INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES,25(18),16.
MLA	Xu, Mingce,et al."Functional Divergence in the Affinity and Stability of Non-Canonical Cysteines and Non-Canonical Disulfide Bonds: Insights from a VHH and VNAR Study".INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES 25.18(2024):16.