Heptad Repeat Sequences are Located Adjacent to Hydrophobic Regions in Several Types of Virus Fusion Glycoproteins

Chambers, P. L.; Pringle, C. R.; Easton, Andrew J.

doi:10.1099/0022-1317-71-12-3075

Cited by 272 publications

(221 citation statements)

References 52 publications

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“…8). The assignment of this internal sequence to the fusion peptide is consistent with the x-ray model, because approximately the same number of residues, 9 and 14 respectively, would separate the last HR1 and HR2 residues visible in the structure from the membrane (12,19). Whereas the x-ray model of the SARS-CoV S2 postfusion core does not reveal the structure assumed by HR1 and HR2 in proximity of the membrane, we obtained some insight into the structure of this region from the crystal structure of a smaller trimeric S2 fragment composed by the shorter peptides N3 (HR1, 914-949) and C2 (HR2, 1149-1193) (Fig.…”

Section: Resultssupporting

confidence: 53%

“…1B). This sequence, in analogy with other viral fusion proteins, has been proposed to be part of a fusion peptide (12,19). Unlike most class I fusion peptides, this sequence is internal rather than at the N terminus to S2.…”

Section: Resultsmentioning

confidence: 99%

“…S1 mediates binding to the receptor which, for the SARS-CoV, has recently been identified as the angiotensin-converting enzyme 2 (10,11). Analysis of CoV S2 sequences shows the presence of two 4-3 heptad repeats (HRs) of hydrophobic residues, HR1 and HR2, typical of coiled coils (12), separated by an Ϸ170-aa-long intervening domain ( Fig. 1) (13).…”

Section: Our Structure Positions a Previously Proposed Internal Fusiomentioning

confidence: 99%

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Structure of a proteolytically resistant core from the severe acute respiratory syndrome coronavirus S2 fusion protein

Supekar¹,

Bruckmann²,

Ingallinella³

et al. 2004

Proc. Natl. Acad. Sci. U.S.A.

116

111

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A coronavirus (CoV) has recently been identified as the causative agent of the severe acute respiratory syndrome (SARS) in humans. CoVs enter target cells through fusion of viral and cellular membranes mediated by the viral envelope glycoprotein S. We have determined by x-ray crystallography the structure of a proteolytically stable core fragment from the heptad repeat (HR) regions HR1 and HR2 of the SARS-CoV S protein. We have also determined the structure of an HR1-HR2 S core fragment, containing a shorter HR1 peptide and a C-terminally longer HR2 peptide that extends up to the transmembrane region. In these structures, three HR1 helices form a parallel coiled-coil trimer, whereas three HR2 peptides pack in an oblique and antiparallel fashion into the coiled-coil hydrophobic grooves, adopting mixed extended and ␣-helical conformations as in postfusion paramyxoviruses F proteins structures. coiled-coil ͉ membrane fusion ͉ viral entry ͉ heptad repeat

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Section: Resultssupporting

confidence: 53%

Section: Resultsmentioning

confidence: 99%

Section: Our Structure Positions a Previously Proposed Internal Fusiomentioning

confidence: 99%

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Structure of a proteolytically resistant core from the severe acute respiratory syndrome coronavirus S2 fusion protein

Supekar¹,

Bruckmann²,

Ingallinella³

et al. 2004

Proc. Natl. Acad. Sci. U.S.A.

116

111

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“…Careful inspection of this sequence reveals that it has characteristics of a class II fusion peptide, notably the conserved proline residue (residue 879) and the high alanine and glycine content. This region has been noted earlier in a theoretical analysis of viral fusion proteins by Chambers et al (32), but was dismissed later when mutation studies of the MHV spike protein pointed to another sequence (residues 971-989) as the likely fusion peptide (33). As we know now, this latter sequence is part of the HR1 domain, which explains the mutation results.…”

Section: Discussionmentioning

confidence: 60%

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

Bosch

Martina

Zee

et al. 2004

Proc. Natl. Acad. Sci. U.S.A.

370

432

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The coronavirus SARS-CoV is the primary cause of the life-threatening severe acute respiratory syndrome (SARS). With the aim of developing therapeutic agents, we have tested peptides derived from the membrane-proximal (HR2) and membrane-distal (HR1) heptad repeat region of the spike protein as inhibitors of SARS-CoV infection of Vero cells. It appeared that HR2 peptides, but not HR1 peptides, were inhibitory. Their efficacy was, however, significantly lower than that of corresponding HR2 peptides of the murine coronavirus mouse hepatitis virus (MHV) in inhibiting MHV infection. Biochemical and electron microscopical analyses showed that, when mixed, SARS-CoV HR1 and HR2 peptides assemble into a six-helix bundle consisting of HR1 as a central triple-stranded coiled coil in association with three HR2 ␣-helices oriented in an antiparallel manner. The stability of this complex, as measured by its resistance to heat dissociation, appeared to be much lower than that of the corresponding MHV complex, which may explain the different inhibitory potencies of the HR2 peptides. Analogous to other class I viral fusion proteins, the six-helix complex supposedly represents a postfusion conformation that is formed after insertion of the fusion peptide, proposed here for coronaviruses to be located immediately upstream of HR1, into the target membrane. The resulting close apposition of fusion peptide and spike transmembrane domain facilitates membrane fusion. The inhibitory potency of the SARS-CoV HR2-peptides provides an attractive basis for the development of a therapeutic drug for SARS.

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“…Hydrophobic 4-3 heptad repeats, characteristic of coiledcoil sequences, are frequently observed within the extracellular domains of viral envelope proteins that mediate fusion of the envelope with cell surface or endosomal membranes during the infectious process (Buckland and Wild, 1989). Examples of this motif are encoded in the human immunodeficiency virus (HIV) GP41, paramyxovirus F, Ebola GP, and baculovirus GP64 membrane proteins (Blissard and Rohrmann, 1989;Whitford et al, 1989;Chambers et al, 1990;Sanchez et al, 1996;Weissenhorn et al, 1998). X-ray crystallographic studies on a number of viral fusion proteins have revealed a common theme of a central trimeric amino-terminal core sequence stabilized by coiled-coil packing, with an antiparallel packing of a second ␣ helix against this central core.…”

mentioning

confidence: 99%

A Discrete Stage of Baculovirus GP64-mediated Membrane Fusion

Kingsley

Behbahani

Rashtian

et al. 1999

MBoC

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Viral fusion protein trimers can play a critical role in limiting lipids in membrane fusion. Because the trimeric oligomer of many viral fusion proteins is often stabilized by hydrophobic 4-3 heptad repeats, higher-order oligomers might be stabilized by similar sequences. There is a hydrophobic 4-3 heptad repeat contiguous to a putative oligomerization domain of Autographa californica multicapsid nucleopolyhedrovirus envelope glycoprotein GP64. We performed mutagenesis and peptide inhibition studies to determine if this sequence might play a role in catalysis of membrane fusion. First, leucine-to-alanine mutants within and flanking the amino terminus of the hydrophobic 4-3 heptad repeat motif that oligomerize into trimers and traffic to insect Sf9 cell surfaces were identified. These mutants retained their wild-type conformation at neutral pH and changed conformation in acidic conditions, as judged by the reactivity of a conformationally sensitive mAb. These mutants, however, were defective for membrane fusion. Second, a peptide encoding the portion flanking the GP64 hydrophobic 4-3 heptad repeat was synthesized. Adding peptide led to inhibition of membrane fusion, which occurred only when the peptide was present during low pH application. The presence of peptide during low pH application did not prevent low pH-induced conformational changes, as determined by the loss of a conformationally sensitive epitope. In control experiments, a peptide of identical composition but different sequence, or a peptide encoding a portion of the Ebola GP heptad motif, had no effect on GP64-mediated fusion. Furthermore, when the hemagglutinin (X31 strain) fusion protein of influenza was functionally expressed in Sf9 cells, no effect on hemagglutinin-mediated fusion was observed, suggesting that the peptide does not exert nonspecific effects on other fusion proteins or cell membranes. Collectively, these studies suggest that the specific peptide sequences of GP64 that are adjacent to and include portions of the hydrophobic 4-3 heptad repeat play a dynamic role in membrane fusion at a stage that is downstream of the initiation of protein conformational changes but upstream of lipid mixing. INTRODUCTIONThe mechanism of membrane fusion is currently a field of intense investigation, both for intracellular trafficking and enveloped viral infection. For viral fusion protein function, we have previously suggested that fusion protein oligomers form transient and cooperative higher-order structures or rings . These transient ring structures are proposed to facilitate conversion of kinetic energy, released during protein conformational change, into work on the target and host membranes required for membrane fusion. Indeed, biochemical evidence for baculovirus GP64 aggregates during membrane fusion has recently been reported (Markovic et al. 1998). Whether and how viral fusion proteins might function in a coordinated and cooperative manner is a major question in the field of membrane fusion. Because hydrophobic 4-3 heptad repeats are observed...

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Heptad Repeat Sequences are Located Adjacent to Hydrophobic Regions in Several Types of Virus Fusion Glycoproteins

Cited by 272 publications

References 52 publications

Structure of a proteolytically resistant core from the severe acute respiratory syndrome coronavirus S2 fusion protein

Structure of a proteolytically resistant core from the severe acute respiratory syndrome coronavirus S2 fusion protein

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

A Discrete Stage of Baculovirus GP64-mediated Membrane Fusion

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