Structure and inhibition of the SARS coronavirus envelope protein ion channel

PLoS Pathog. 2009 Jul;5(7):e1000511. doi: 10.1371/journal.ppat.1000511. Epub 2009 Jul 10.

Abstract

The envelope (E) protein from coronaviruses is a small polypeptide that contains at least one alpha-helical transmembrane domain. Absence, or inactivation, of E protein results in attenuated viruses, due to alterations in either virion morphology or tropism. Apart from its morphogenetic properties, protein E has been reported to have membrane permeabilizing activity. Further, the drug hexamethylene amiloride (HMA), but not amiloride, inhibited in vitro ion channel activity of some synthetic coronavirus E proteins, and also viral replication. We have previously shown for the coronavirus species responsible for severe acute respiratory syndrome (SARS-CoV) that the transmembrane domain of E protein (ETM) forms pentameric alpha-helical bundles that are likely responsible for the observed channel activity. Herein, using solution NMR in dodecylphosphatidylcholine micelles and energy minimization, we have obtained a model of this channel which features regular alpha-helices that form a pentameric left-handed parallel bundle. The drug HMA was found to bind inside the lumen of the channel, at both the C-terminal and the N-terminal openings, and, in contrast to amiloride, induced additional chemical shifts in ETM. Full length SARS-CoV E displayed channel activity when transiently expressed in human embryonic kidney 293 (HEK-293) cells in a whole-cell patch clamp set-up. This activity was significantly reduced by hexamethylene amiloride (HMA), but not by amiloride. The channel structure presented herein provides a possible rationale for inhibition, and a platform for future structure-based drug design of this potential pharmacological target.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amiloride / analogs & derivatives
  • Amiloride / metabolism
  • Amiloride / pharmacology
  • Cell Line
  • Humans
  • Ion Channels / antagonists & inhibitors*
  • Ion Channels / chemistry*
  • Ion Channels / metabolism
  • Models, Molecular
  • Nuclear Magnetic Resonance, Biomolecular
  • Patch-Clamp Techniques
  • Protein Binding
  • Protein Conformation
  • Protein Stability
  • Protein Structure, Tertiary
  • Reproducibility of Results
  • SARS Virus / chemistry*
  • SARS Virus / metabolism
  • Viral Envelope Proteins / antagonists & inhibitors*
  • Viral Envelope Proteins / chemistry*
  • Viral Envelope Proteins / metabolism

Substances

  • Ion Channels
  • Viral Envelope Proteins
  • 5-(N,N-hexamethylene)amiloride
  • Amiloride