Structure and inhibition of the SARS coronavirus envelope protein ion channel.
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.
Pervushin
Konstantin
K
School of Biological Sciences, Nanyang Technological University, Singapore. kpervushin@ntu.edu.sg
Tan
Edward
E
Parthasarathy
Krupakar
K
Lin
Xin
X
Jiang
Feng Li
FL
Yu
Dejie
D
Vararattanavech
Ardcharaporn
A
Soong
Tuck Wah
TW
Liu
Ding Xiang
DX
Torres
Jaume
J
D000584:Amiloride / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology; D002460:Cell Line; D006801:Humans; D007473:Ion Channels / Q000037:antagonists & inhibitors* / Q000737:chemistry* / Q000378:metabolism; D008958:Models, Molecular; D019906:Nuclear Magnetic Resonance, Biomolecular; D018408:Patch-Clamp Techniques; D011485:Protein Binding; D011487:Protein Conformation; D055550:Protein Stability; D017434:Protein Structure, Tertiary; D015203:Reproducibility of Results; D045473:Severe acute respiratory syndrome-related coronavirus / Q000737:chemistry* / Q000378:metabolism; D014759:Viral Envelope Proteins / Q000037:antagonists & inhibitors* / Q000737:chemistry* / Q000378:metabolism
D007473:Ion Channels; D014759:Viral Envelope Proteins; C057360:5-(N,N-hexamethylene)amiloride; D000584:Amiloride