Structure and inhibition of the SARS coronavirus envelope protein ion channel

• Published in: PLoS pathogens Vol. 5; no. 7; p. e1000511
• Main Authors: Pervushin, Konstantin, Tan, Edward, Parthasarathy, Krupakar, Lin, Xin, Jiang, Feng Li, Yu, Dejie, Vararattanavech, Ardcharaporn, Soong, Tuck Wah, Liu, Ding Xiang, Torres, Jaume
• Format: Journal Article
• Language: English
• Published: United States Public Library of Science 01.07.2009; Public Library of Science (PLoS)
• Subjects: Amiloride - analogs & derivatives; Amiloride - metabolism; Amiloride - pharmacology; Binding sites; Biomedical research; Biophysics; Biophysics/Experimental Biophysical Methods; Biophysics/Membrane Proteins and Energy Transduction; Biophysics/Structural Genomics; Cell Line; Clinical trials; Coronaviruses; Development and progression; Genetic aspects; Humans; Ion channels; Ion Channels - antagonists & inhibitors; Ion Channels - chemistry; Ion Channels - metabolism; Mammals; Models, Molecular; Nuclear Magnetic Resonance, Biomolecular; Pandemics; Patch-Clamp Techniques; Physiological aspects; Polypeptides; Protein Binding; Protein Conformation; Protein Stability; Protein Structure, Tertiary; Proteins; Reproducibility of Results; SARS coronavirus; SARS Virus - chemistry; SARS Virus - metabolism; Severe acute respiratory syndrome; Viral Envelope Proteins - antagonists & inhibitors; Viral Envelope Proteins - chemistry; Viral Envelope Proteins - metabolism; Viral proteins; Singapore
• ISSN: 1553-7374; 1553-7366; 1553-7374
• DOI: 10.1371/journal.ppat.1000511

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.