Crystal structure of NL63 respiratory coronavirus receptor-binding domain complexed with its human receptor - PubMed (original) (raw)

Crystal structure of NL63 respiratory coronavirus receptor-binding domain complexed with its human receptor

Kailang Wu et al. Proc Natl Acad Sci U S A. 2009.

Abstract

NL63 coronavirus (NL63-CoV), a prevalent human respiratory virus, is the only group I coronavirus known to use angiotensin-converting enzyme 2 (ACE2) as its receptor. Incidentally, ACE2 is also used by group II SARS coronavirus (SARS-CoV). We investigated how different groups of coronaviruses recognize the same receptor, whereas homologous group I coronaviruses recognize different receptors. We determined the crystal structure of NL63-CoV spike protein receptor-binding domain (RBD) complexed with human ACE2. NL63-CoV RBD has a novel beta-sandwich core structure consisting of 2 layers of beta-sheets, presenting 3 discontinuous receptor-binding motifs (RBMs) to bind ACE2. NL63-CoV and SARS-CoV have no structural homology in RBD cores or RBMs; yet the 2 viruses recognize common ACE2 regions, largely because of a "virus-binding hotspot" on ACE2. Among group I coronaviruses, RBD cores are conserved but RBMs are variable, explaining how these viruses recognize different receptors. These results provide a structural basis for understanding viral evolution and virus-receptor interactions.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Fig. 1.

Fig. 1.

Structure of NL63-CoV RBD complexed with human ACE2. (A) Domain structure of the NL63-CoV spike protein. Unique, unique region; NTR, N-terminal region; Central, central region; CTR, C-terminal region; HR-N, heptad-repeat N; HR-C, heptad-repeat C; TM, transmembrane anchor; IC, intracellular tail. The unique domain only exists in NL63-CoV and is not involved in receptor binding (19, 20). (B) Overall structure of NL63-CoV RBD complexed with human ACE2. The RBD core is in cyan, RBMs in red, and ACE2 in green. (C) Averaged electron density map contoured at 1.0 σ and covering a portion of the NL63-CoV–ACE2 interface. (D) Sequence and secondary structures of NL63-CoV RBD. Beta-strands are drawn as arrows. RBMs are in red; the remainder of the RBD is in cyan. Disordered regions are shown as dashed lines. (E) Kinetics and binding affinity of NL63-CoV RBD and human ACE2 by surface plasmon resonance using Biacore. Structural illustrations were made using Povscript (31).

Fig. 2.

Fig. 2.

Structural comparison of NL63-CoV and SARS-CoV RBDs. (A) Domain structure of NL63-CoV S1. The boundaries of the RBD were determined by limited proteolysis of longer S1 fragments, followed by N-terminal sequencing and mass spectrometric analysis of the digestion fragments. The RBMs were identified from the crystal structure of the NL63-CoV RBD in complex with ACE2. (B) Domain structure of SARS-CoV S1 (5). (C) Another view of the structure of the NL63-CoV-RBD–ACE2 complex, which is derived by rotating the one in Fig. 1_B_ by 90° clockwise along a vertical axis. (D) Structure of the SARS-CoV-RBD–ACE2 complex (PDB 2AJF) (5), from the same orientation as in C. (E) Schematic illustration of the topology of NL63-CoV RBD. Strands are drawn as arrows. (F) Schematic illustration of the topology of SARS-CoV RBD.

Fig. 3.

Fig. 3.

Structural features of NL63-CoV RBD and RBMs. (A) Beta-sandwich core structure of NL63-CoV RBD. Hydrophobic residues between β-sheet layers are in yellow, cysteines in blue, glycans in green, and glycosylated asparagines in magenta. There exist 3 predicted N-linked glycosylation sites in the RBD (Asn-486, Asn-506, and Asn-512), and 2 of them are confirmed in the structure (Asn-486 and Asn-512). (B) NL63-CoV RBMs. (C) Residues on NL63-CoV RBMs that directly contact ACE2.

Fig. 4.

Fig. 4.

Structural comparison of NL63-CoV–ACE2 and SARS-CoV–ACE2 interfaces. (A) Enlarged view of the NL63-CoV–ACE2 interface, from the same orientation as in Fig. 2_C_. VBMs on ACE2 are in blue, and RBMs on NL63-CoV are in red. Arrow indicates the bowl-shaped cavity surrounded by 3 viral RBMs. (B) Enlarged view of the SARS-CoV–ACE2 interface, from the same orientation as in A. (C) Footprint of NL63-CoV on the surface of ACE2. The view is derived from the one in A by rotating ACE2 by 90° along a horizontal axis, in such a way that the edge facing the viewer moves up. VBM1 residues are in orange, VBM2 residues in magenta, and VBM3 residues in red. (D) Footprint of SARS-CoV on the surface of ACE2, from the same orientation as in C. VBM1b residues are in green.

Fig. 5.

Fig. 5.

A common virus-binding hotspot on ACE2 for the binding of both NL63-CoV and SARS-CoV. (A) Stick-and-ball representation of the hotspot at the NL63-CoV–ACE2 interface. (B) Corey-Pauling-Koltun representation of the hotspot at the NL63-CoV–ACE2 interface. (C) Stick-and-ball representation of the hotspot at the SARS-CoV–ACE2 interface. (D) Corey-Pauling-Koltun representation of the hotspot at the SARS-CoV–ACE2 interface.

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