Solution structure of the cytoplasmic domain of NhaP2 a K+/H+ antiporter from Vibrio cholera

Biochim Biophys Acta Biomembr. 2020 Jun 1;1862(6):183225. doi: 10.1016/j.bbamem.2020.183225. Epub 2020 Feb 29.

Abstract

NhaP2 is a K+/H+ antiporter from Vibrio cholerae which consists of a transmembrane domain and a cytoplasmic domain of approximately 200 amino acids, both of which are required for cholera infectivity. Here we present the solution structure for a 165 amino acid minimal cytoplasmic domain (P2MIN) form of the protein. The structure reveals a compact N-terminal domain which resembles a Regulator of Conductance of K+ channels (RCK) domain connected to a more open C-terminal domain via a flexible 20 amino acid linker. NMR titration experiments showed that the protein binds ATP through its N-terminal domain, which was further supported by waterLOGSY and Saturation Transfer Difference NMR experiments. The two-domain organisation of the protein was confirmed by BIOSAXS, which also revealed that there are no detectable-ATP-induced conformational changes in the protein structure. Finally, in contrast to all known RCK domain structures solved to date, the current work shows that the protein is a monomer.

Keywords: Antiporter; C-terminal tail and 3D structure; Integrated approach in structural biophysics; RCK domains.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenosine Triphosphate / metabolism
  • Antiporters / chemistry
  • Antiporters / metabolism
  • Bacterial Proteins / chemistry*
  • Bacterial Proteins / metabolism
  • Binding Sites
  • Cytoplasm / chemistry
  • Nuclear Magnetic Resonance, Biomolecular
  • Potassium-Hydrogen Antiporters / chemistry*
  • Potassium-Hydrogen Antiporters / metabolism
  • Protein Conformation
  • Protein Domains*
  • Vibrio cholerae / chemistry*

Substances

  • Antiporters
  • Bacterial Proteins
  • Potassium-Hydrogen Antiporters
  • Adenosine Triphosphate