Crystal structure of severe acute respiratory syndrome coronavirus spike protein fusion core

Abstract

Severe acute respiratory syndrome coronavirus is a newly emergent virus responsible for a recent outbreak of an atypical pneumonia. The coronavirus spike protein, an enveloped glycoprotein essential for viral entry, belongs to the class I fusion proteins and is characterized by the presence of two heptad repeat (HR) regions, HR1 and HR2. These two regions are understood to form a fusion-active conformation similar to those of other typical viral fusion proteins. This hairpin structure likely juxtaposes the viral and cellular membranes, thus facilitating membrane fusion and subsequent viral entry. The fusion core protein of severe acute respiratory syndrome coronavirus spike protein was crystallized, and the structure was determined at 2.8 A of resolution. The fusion core is a six-helix bundle with three HR2 helices packed against the hydrophobic grooves on the surface of central coiled coil formed by three parallel HR1 helices in an oblique antiparallel manner. This structure shares significant similarity with the fusion core structure of mouse hepatitis virus spike protein and other viral fusion proteins, suggesting a conserved mechanism of membrane fusion. Drug discovery strategies aimed at inhibiting viral entry by blocking hairpin formation, which have been successfully used in human immunodeficiency virus 1 inhibitor development, may be applicable to the inhibition of severe acute respiratory syndrome coronavirus on the basis of structural information provided here. The relatively deep grooves on the surface of the central coiled coil will be a good target site for the design of viral fusion inhibitors.

Fig. 1

Structure determination of the MHV spike protein fusion core trimer.A, schematic diagram of SARS-CoV spike protein indicating the location of structurally significant domains. S1 and S2 are formed after proteolytic cleavage (vertical arrow) and noncovalently linked. The enveloped protein has an N-terminal signal sequence (SS) and a transmembrane domain (TM) adjacent to the C terminus. S2 contains two HR (heptad repeat) regions (hatched bars), HR1 and HR2 as indicated. The HR1 (898–1005) and HR2 (1145–1184) used in this study were derived from the LearnCoil-VMF prediction program (37). The 2-Helix protein construct consists of HR2 and part of HR1, which is the major region binding HR2, connected by a 22-amino acid linker (LVPRGSGGSGGSGGLEVLFQGP) as indicated. B, sequence alignment of coronavirus spike protein HR1 and HR2 regions. Letters above the sequence indicate the predicted hydrophobic residues at the a and d positions in two heptad repeat regions, which are highly conserved. FP, feline panleukopenia; IBV, infectious bronchitis virus; FIPV, feline infectious peritonitis virus.

Fig. 2

Overall views of the fusion core structure.A, top view of the SARS-CoV spike protein fusion core structure showing the 3-fold axis of the trimer. B, side view of the SARS-CoV spike protein fusion core structure showing the six-helix bundle.

Fig. 4

Comparison between fusion core structure of SARS-CoV and MHV.A, side view showing a structural comparison between SARS-CoV spike protein fusion core (colored in green) and MHV spike protein fusion core (colored in purple). The columns at both sides of the map represent two HR1 and HR2 regions of MHV and SARS fusion cores. The numbers at the end of these columns represent the specific boundaries of the HR1-HR2 interaction region in the two structures. B, surface map showing the comparison between hydrophobic grooves on the surface of three central HR1 regions of MHV (left side) and SARS-CoV (right side). The figure on the right side shows the deep and relatively shallow grooves on the surface of central HR1 coiled coil of SARS-CoV. Three numbers, 1–3, in circles represent three deep cavities, composing the deep grooves. The residues represent the boundaries of the grooves and cavities.

Fig. 3

Detailed structure of the SARS-CoV spike protein fusion core and OXO motifs in HR2 regions.A, surface map showing the hydrophobic grooves on the surface of the central coiled coil (right side). Three HR2 helices pack against the hydrophobic grooves in an oblique antiparallel manner (left side). The helical regions and extended regions in HR2 helices could be observed clearly, and the boundaries of these regions are marked. B, OXO motifs in HR2 regions of SARS-CoV spike protein fusion core structure. The enlarged images show two regions containing OXO motifs. The hydrophobic residues in these motifs all pack against the hydrophobic grooves on the surface of three HR1 helices.