A Core Trimer of the Paramyxovirus Fusion Protein: Parallels to Influenza Virus Hemagglutinin and HIV-1 gp41

Abstract

The paramyxovirus fusion (F) protein mediates membrane fusion. The biologically active F protein consists of a membrane distal subunit, F2, and a membrane-anchored subunit, F1. We have identified a highly stable structure composed of peptides derived from the F1 heptad repeat A, which abuts the hydrophobic fusion peptide (peptide N-1), and the F1 heptad repeat B, located 270 residues downstream and adjacent to the transmembrane domain (peptides C-1 and C-2). In isolation, peptide N-1 is 47% alpha-helical and peptide C-1 and C-2 are unfolded. When mixed together, peptides N1 + C1 form a thermostable (Tm >90 degreesC), 82% alpha-helical, discrete trimer of heterodimers (mass 31,300 Mr) that is resistant to denaturation by 2% SDS at 40 degreesC. We suggest that this alpha-helical trimeric complex represents the core most stable form of the F protein that either is fusion competent or forms after fusion has occurred. Peptide C-1 is a potent inhibitor of both the lipid mixing and the aqueous content mixing fusion activity of the SV5 F protein. In contrast, peptides N-1 and N-2 inhibit cytoplasmic content mixing but not lipid mixing, leading to a stable hemifusion state. Thus, these peptides define functionally different steps in the fusion process. The parallels among both the fusion processes and the protein structures of paramyxovirus F proteins, HIV gp41, and influenza virus hemagglutinin are discussed, as the analogies are indicative of a conserved paradigm for fusion promotion among fusion proteins from widely disparate viruses.

Severe acute respiratory syndrome coronavirus (SARS-CoV) infection inhibition using spike protein heptad repeat-derived peptides

“…Peptides corresponding to the HR1 and HR2 domains were found to associate tightly into a complex, as has been observed previously for retrovirus, paramyxovirus, and coronavirus fusion proteins (2,12,23,24). Analogous to the HIV-1 gp41, SV5 F, and HRSV F proteins (25)(26)(27)(28), the HR1-HR2 complex was found to consist of a six-helix bundle that is composed of three HR1 and three HR2 ␣-helical peptides.…”

“…Under the electron microscope, the complexes were visible as rod-like structures similar in appearance and dimensions as the HR1-HR2 complexes. The formation of a protease-resistant homotrimeric coiled coil by the isolated HR1 peptides has been reported for the SV5 F protein (12) and for Moloney murine leukemia virus gp41 (29), but not for HIV-1 or SIV gp41 (23,24), indicating the variation in stability of the HR1 core trimer among class I viral fusion proteins. There are no indications that a homotrimeric interaction between coronavirus HR1 regions already occurs in the native virion, i.e., before binding to the cell receptor.…”

“…As has been described for retrovirus, paramyxovirus, and coronavirus fusion proteins (2,(11)(12)(13)(14)(15)(16), peptides derived from the HR2 domain can inhibit virus infection, most likely by interfering with six-helix bundle formation, a process essential to drive the membrane fusion reaction and, thus, to initiate infection. For HIV-1, this has led to the development of peptide inhibitors, one of which has recently been licensed as a drug against AIDS.…”

A Discrete Stage of Baculovirus GP64-mediated Membrane Fusion

“…Specifically for HIV gp41, it has been shown that two separate heptad repeats form a six-helix bundle with the amino-terminal heptad repeats of gp41 to create a central trimeric core, with the carboxy-terminal heptad repeat packing in an antiparallel manner within the outside grooves of the core trimer (Weissenhorn et al, 1996(Weissenhorn et al, , 1997Chan et al, 1997). This general architecture is observed in other viral fusion proteins, such as other retroviruses Fass and Kim, 1995;Lu et al, 1995;Fass et al, 1996;Malashkevich et al, 1998), Ebola GP (Wiessenhorn et al, 1998Malashkevich et al, 1999), paramyxovirus F (Joshi et al, 1998;Baker et al, 1999;Dutch et al, 1999), and influenza hemagglutinin (HA) proteins (Carr and Kim, 1993;Bullough et al, 1994). Conversion of fusion proteins to a rod-like structure via formation of a long trimeric coiled coil upon low pH application has been observed and forms the basis of a proposed spring-loaded mechanism by which hydrophobic amino-terminal "fusion peptides" are projected into the target membrane, ultimately resulting in fusion of the viral envelope with the target cell membrane (Carr and Kim 1993;Chan et al, 1997;Weissenhorn et al, 1997Weissenhorn et al, , 1999.…”

“…For HIV gp41 peptide inhibition, it has been predicted that peptides would exert inhibitory effects during the conformational change to the fusion-active conformation (Wiessenhorn et al, 1997). In fact, recent work on peptide inhibition of HIV gp41 and parainfluenza also suggests that peptide inhibition may function dynamically by preventing the hypothesized collapse of the extended coiledcoil structure back on itself as part of a proposed mechanism by which target and host membranes are merged (Joshi et al, 1998;Munoz-Barroso et al, 1998;Baker et al, 1999). Also, the HIV inhibitory peptide binds to gp41 only after gp120 interacts with its receptor(s) and gp41 is activated (Furata et al, 1998).…”