Nanobodies as negative allosteric modulators for human calcium sensing receptor

Qianqian Cui; Lu Wang; Haonan Wang; Xiaochen Chen; Li Han; Tengjie Geng; Yongjun Kou; Wenqing Zhang; Mei Dai; Huarui Qiao; Zengchao Sun; Lingyun Li; Zhongyun Lan; Hongxi Xu; Jiaqiang Xu; Yuanyuan Dai; Yong Geng

doi:10.1016/j.bbrc.2023.149401

Nanobodies as negative allosteric modulators for human calcium sensing receptor

Biochem Biophys Res Commun. 2024 Feb 5:695:149401. doi: 10.1016/j.bbrc.2023.149401. Epub 2023 Dec 21.

Authors

Qianqian Cui¹, Lu Wang², Haonan Wang², Xiaochen Chen², Li Han², Tengjie Geng², Yongjun Kou², Wenqing Zhang², Mei Dai², Huarui Qiao², Zengchao Sun², Lingyun Li², Zhongyun Lan², Hongxi Xu³, Jiaqiang Xu⁴, Yuanyuan Dai⁵, Yong Geng⁶

Affiliations

¹ NEST Lab, Department of Chemistry, College of Science, Shanghai University, Shanghai, 200444, China; The CAS Key Laboratory of Receptor Research, Stake Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai, 201203, China.
² The CAS Key Laboratory of Receptor Research, Stake Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai, 201203, China.
³ School of Pharmacy, Shanghai University of Traditional Chinese Medicine, Shanghai, 201203, China. Electronic address: daiyuanyuan@cicams.ac.cn.
⁴ NEST Lab, Department of Chemistry, College of Science, Shanghai University, Shanghai, 200444, China. Electronic address: xujiaqiang@shu.edu.cn.
⁵ Department of Pharmacy, National Cancer Center/ National Clinical Research Center for Cancer/Cancer Hospital, and Peking Union Medical College, Chinese Academy of Medical Science, Beijing, 100021, China. Electronic address: xuhongxi88@gmail.com.
⁶ The CAS Key Laboratory of Receptor Research, Stake Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai, 201203, China; University of Chinese Academy of Sciences, Beijing, 100049, China. Electronic address: gengyong@simm.ac.cn.

PMID: 38154264
DOI: 10.1016/j.bbrc.2023.149401

Abstract

Human calcium sensing receptor (CaSR) senses calcium ion concentrations in vivo and is an important class of drug targets. Mutations in the receptor can lead to disorders of calcium homeostasis, including hypercalcemia and hypocalcemia. Here, 127 CaSR-targeted nanobodies were generated from camels, and four nanobodies with inhibitory function were further identified. Among these nanobodies, NB32 can effectively inhibit the mobilization of intracellular calcium ions (Ca²⁺_i) and suppress the G_12/13 and ERK_1/2 signaling pathways downstream of CaSR. Moreover, it enhanced the inhibitory effect of the calcilytics as a negative allosteric modulator (NAM). We determined the structure of complex and found NB32 bound to LB2 (Ligand-binding 2) domain of CaSR to prevent the interaction of LB2 domains of two protomers to stabilize the inactive state of CaSR.

Keywords: Cryo-electron microscope; Extracellular calcium sensing receptor; G-protein-coupled receptor; Nanobody; Systemic Ca(2+) homoeostasis.

MeSH terms

Calcium / metabolism
Humans
Hypercalcemia* / genetics
Hypocalcemia* / genetics
Receptors, Calcium-Sensing / metabolism
Single-Domain Antibodies*

Substances

Receptors, Calcium-Sensing
Calcium
Single-Domain Antibodies