Nanobodies as negative allosteric modulators for human calcium sensing receptor
Qianqian Cui1, Lu Wang2, Haonan Wang2
1NEST 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.
Biochemical and Biophysical Research Communications
|December 28, 2023
Summary
Camel-derived nanobodies targeting the calcium sensing receptor (CaSR) were developed. NB32 nanobody inhibits CaSR activity by stabilizing its inactive state, offering potential for calcium disorder treatments.
Area of Science:
- Biochemistry
- Pharmacology
- Structural Biology
Background:
- The human calcium sensing receptor (CaSR) regulates calcium homeostasis.
- CaSR mutations cause calcium imbalance disorders like hypercalcemia and hypocalcemia.
- CaSR is a significant drug target for treating these disorders.
Purpose of the Study:
- To generate and identify CaSR-specific nanobodies with inhibitory functions.
- To characterize the inhibitory mechanism of a lead nanobody, NB32.
- To elucidate the structural basis of NB32-CaSR interaction.
Main Methods:
- Generation and screening of 127 camel-derived nanobodies against CaSR.
- Functional assays to assess inhibition of intracellular calcium mobilization.
- Analysis of downstream signaling pathways (G12/13, ERK1/2).
- Co-crystallization and structural determination of the CaSR-NB32 complex.
Main Results:
- Four inhibitory nanobodies were identified from 127 candidates.
- NB32 effectively inhibited CaSR-mediated intracellular calcium mobilization.
- NB32 suppressed G12/13 and ERK1/2 signaling and enhanced calcilytic efficacy.
- Structural analysis revealed NB32 binds the CaSR LB2 domain, preventing protomer interaction and stabilizing the inactive receptor.
Conclusions:
- NB32 is a potent CaSR inhibitor that stabilizes the receptor's inactive conformation.
- NB32's mechanism involves allosteric modulation by binding the LB2 domain.
- This nanobody represents a promising therapeutic candidate for calcium-related disorders.
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