NMR structures and localization of the potential fusion peptides and the pre-transmembrane region of SARS-CoV: Implications in membrane fusion

Abstract

Severe acute respiratory syndrome-associated coronavirus (SARS-CoV) poses a serious public health hazard. The S2 subunit of the S glycoprotein of SARS-CoV carries out fusion between the virus and the host cells. However, the exact mechanism of the cell fusion process is not well understood. Current model suggests that a conformational transition, upon receptor recognition, of the two heptad core regions of S2 may expose the hydrophobic fusogenic peptide or fusion peptide for membrane insertion. Three regions of the S2 subunit have been proposed to be involved in cell-cell fusion. The N-terminal fusion peptide (FP, residues 770-788), an internal fusion peptide (IFP, residues 873-888) and the pre-transmembrane region (PTM, residues 1185-1202) demonstrated interactions with model lipid membranes and potentially involved in the fusion process. Here, we have determined atomic resolution structures of these three peptides in DPC detergent micelles by solution NMR. FP assumes α-helical conformation with significant distortion at the central Gly residues; enabling a close packing among sidechains of aromatic residues including W, Y and F. The 3-D structure of PMT is characterized by a helix-loop-helix with extensive aromatic interactions within the helices. IFP adopts a rather straight α-helical conformation defined by packing among sidechains of aromatic and aliphatic residues. Paramagnetic spin labeled NMR has demonstrated surface localization of PMT whereas FP and IFP inserted into the micelles. Collectively, data presented in this study will aid in understanding fusion mechanism of SARS-CoV.

Bat-to-human: spike features determining ‘host jump’ of coronaviruses SARS-CoV, MERS-CoV, and beyond

“…Multiple fusion-related components in SARS-CoV S2 have been extensively studied thus far ( Figure 1A,B). These include the fusion core composed of HR1 and HR2 [27,28] and at least three membranotropic regions that are denoted as the fusion peptide (FP), internal fusion peptide (IFP), and pretransmembrane domain (PTM), respectively [35]. The two HR modules are separately dispatched in S2 and are separated from each other by $200 residues.…”

“…The HR regions are further flanked by the three membranotropic components. Both FP and IFP are located upstream of HR1, spanning residues 770-788 and 873-888, respectively, while PTM is distally downstream of HR2 and directly precedes the transmembrane domain of SARS-CoV S. All of these three components are able to partition into the phospholipid bilayer to disturb membrane integrity [38], and their structural features have recently been elucidated [35]. FP assumes an a-helical conformation but shows significant distortion at its center.…”

“…PTM assumes a helix-loophelix fold. It should be noted that all three components can create a hydrophobic side-surface ( Figure 1B), explaining their bilayer-binding capacities [35]. The exact role of these putative fusion peptides in virus-cell fusion, however, remains to be fully examined; for example, it is currently unknown whether FP, IFP, and PTM function individually or in a synergistic manner.…”

“…A portion of the interstrand loop extends extensively over the surface of the core subdomain, and, together with the two b-strands, anchors the external region to the core like a clamp ( Figure 1B). It is interesting that one structure of the free SARS-CoV RBD unexpectedly revealed the possible dimerization of the protein through The individual components of S that were either experimentally characterized in previous studies -including receptor-binding domain (RBD), fusion peptide (FP), internal fusion peptide (IFP), heptad repeat 1/2 (HR1/2), and pretransmembrane domain (PTM) [13,27,35] -or are based on bioinformatics analyses, for example, N-terminal domain (NTD), are marked with the boundary-residue numbers listed below. The S1/S2 cleavage sites and the S2'-recognition site are highlighted.…”

Membranotropic peptides mediating viral entry

“…Indeed, peptides corresponding to the pre‐TM region partition into membrane interfaces and likely cooperate with fusion peptides and TM domains during apposition of membranes, enhancing the overall hydrophobicity of the environment and contributing to the distortion of the lipid membranes required for fusion . The pre‐TM of HSV‐1 gH interacts strongly with membranes; the pre‐TM of Gp47 of foamy virus induces fusion of model membranes; the aromatic domain of the glycoprotein S2 of severe acute respiratory syndrome virus (SARS) partitions into lipid membranes and perturbs their integrity; the pre‐TM region of the GP2 protein from Ebola promotes perturbations of membranes when in a helical structure …”