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

Elife. 2021 Jul 28:10:e67115. doi: 10.7554/eLife.67115.

Abstract

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.

Keywords: ASIC1; MitTx antagonism; Pctx1 potentiation; cryo-electron microscopy; molecular biophysics; nanobody; structural biology.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Acid Sensing Ion Channels / chemistry*
  • Acid Sensing Ion Channels / ultrastructure
  • Animals
  • Camelids, New World
  • Cryoelectron Microscopy
  • Protein Binding
  • Single-Domain Antibodies / chemistry*
  • Single-Domain Antibodies / ultrastructure

Substances

  • ASIC1 protein, human
  • Acid Sensing Ion Channels
  • Single-Domain Antibodies