Structure and analysis of nanobody binding to the human ASIC1a ion channel

• Published in: eLife Vol. 10
• Main Authors: Wu, Yangyu, Chen, Zhuyuan, Sigworth, Fred J, Canessa, Cecilia M
• Format: Journal Article
• Language: English
• Published: England eLife Science Publications, Ltd 28.07.2021; eLife Sciences Publications Ltd; eLife Sciences Publications, Ltd
• Subjects: Acid Sensing Ion Channels - chemistry; Acid Sensing Ion Channels - ultrastructure; Addictions; Amino acids; Analgesics; Analysis; Animals; Antibodies; ASIC1; Binding sites; Camelids, New World; Cloning; cryo-electron microscopy; Cryoelectron Microscopy; Electron microscopy; Genomes; Immunization; Immunoglobulins; Ischemia; Lymphocytes; MitTx antagonism; Nanobodies; nanobody; Pain; Pctx1 potentiation; Phylogenetics; Poisonous snakes; Protein Binding; Proteins; Single-Domain Antibodies - chemistry; Single-Domain Antibodies - ultrastructure; Structural Biology and Molecular Biophysics; Therapeutic applications; Toxins; Venom; Canada; MitTx antagonism; structural biology; nanobody; molecular biophysics; Pctx1 potentiation; ASIC1; cryo-electron microscopy
• ISSN: 2050-084X; 2050-084X
• DOI: 10.7554/eLife.67115

ASIC1a is a proton-gated sodium channel involved in modulation of pain, fear, addiction, and ischemia-induced neuronal injury. We report isolation and characterization of alpaca-derived nanobodies (Nbs) that specifically target human ASIC1a. Cryo-electron microscopy of the human ASIC1a channel at pH 7.4 in complex with one of these, Nb.C1, yielded a structure at 2.9 Å resolution. It is revealed that Nb.C1 binds to a site overlapping with that of the Texas coral snake toxin (MitTx1) and the black mamba venom Mambalgin-1; however, the Nb.C1-binding site does not overlap with that of the inhibitory tarantula toxin psalmotoxin-1 (PcTx1). Fusion of Nb.C1 with PcTx1 in a single polypeptide markedly enhances the potency of PcTx1, whereas competition of Nb.C1 and MitTx1 for binding reduces channel activation by the toxin. Thus, Nb.C1 is a molecular tool for biochemical and structural studies of hASIC1a; a potential antidote to the pain-inducing component of coral snake bite; and a candidate to potentiate PcTx1-mediated inhibition of hASIC1a in vivo for therapeutic applications.