Mutations in the fusion peptide and heptad repeat regions of the Newcastle disease virus fusion protein block fusion

Sergel-Germano, T; McQuain, Cathy; Morrison, Trudy G.

doi:10.1128/jvi.68.11.7654-7658.1994

Cited by 89 publications

(46 citation statements)

References 28 publications

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“…As has been well documented for the paramyxovirus F protein and other class I viral fusion proteins, nonpolar residues at the a and d positions in the HRA region are important in the formation of the coiled-coil hairpin (18,19,31,55). Mutations to individual a and d residues in HRA also affect the folding and fusogenic activity of intact NDV and PIV5 F proteins (51,53,60), a finding that suggests that these residues are important in the formation and activation of the native conformation of the F protein. Many other HRA residues, in addition to a and d residues, are involved in structural interactions that maintain the prefusion conformation of the F protein (64).…”

Section: Discussionmentioning

confidence: 93%

“…Many of the conservative mutations to HRA residues reduce expression of the SeV F protein. Mutation of nonpolar "a" and "d" residues in HRA of the NDV F protein to lysine residues decreases cell surface expression and eliminates membrane fusion activity (51,53). In contrast, mutation of a and d residues in HRA of the PIV5 F protein to methionine residues does not, in many cases, disrupt the ␣-helix formation, protein expression and processing, or membrane fusion activity (60).…”

Section: Resultsmentioning

confidence: 99%

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Spring-Loaded Heptad Repeat Residues Regulate the Expression and Activation of Paramyxovirus Fusion Protein

Luque

Russell

2007

J Virol

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During viral entry, the paramyxovirus fusion (F) protein fuses the viral envelope to a cellular membrane. Similar to other class I viral fusion glycoproteins, the F protein has two heptad repeat regions (HRA and HRB) that are important in membrane fusion and can be targeted by antiviral inhibitors. Upon activation of the F protein, HRA refolds from a spring-loaded, crumpled structure into a coiled coil that inserts a hydrophobic fusion peptide into the target membrane and binds to the HRB helices to form a fusogenic hairpin. To investigate how F protein conformational changes are regulated, we mutated in the Sendai virus F protein a highly conserved 10-residue sequence in HRA that undergoes major structural changes during protein refolding. Nine of the 15 mutations studied caused significant defects in F protein expression, processing, and fusogenicity. Conversely, the remaining six mutations enhanced the fusogenicity of the F protein, most likely by helping spring the HRA coil. Two of the residues that were neither located at "a" or "d" positions in the heptad repeat nor conserved among the paramyxoviruses were key regulators of the folding and fusion activity of the F protein, showing that residues not expected to be important in coiled-coil formation may play important roles in regulating membrane fusion. Overall, the data support the hypothesis that regions in the F protein that undergo dramatic changes in secondary and tertiary structure between the prefusion and hairpin conformations regulate F protein expression and activation.

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Section: Discussionmentioning

confidence: 93%

Section: Resultsmentioning

confidence: 99%

Spring-Loaded Heptad Repeat Residues Regulate the Expression and Activation of Paramyxovirus Fusion Protein

Luque

Russell

2007

J Virol

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“…Those include sequence conservation among coronaviruses, hydrophobicity, richness in alanine and glycine residues, and the ability to be modeled into a sided helix and bounded by charged residues. Studies of heptad repeat regions in many viral fusion proteins have revealed that they are also essential for membrane fusion (Buckland et al, 1992;Dubay et al, 1992;Sergel-Germano et al, 1994). Although fusion peptides are generally found adjacent to heptad repeats but not within the heptad repeat, there is no indication that fusion peptides cannot reside in a heptad repeat.…”

Section: Discussionmentioning

confidence: 99%

“…Although fusion peptides are generally found adjacent to heptad repeats but not within the heptad repeat, there is no indication that fusion peptides cannot reside in a heptad repeat. It is postulated that heptad repeats may also be able to insert into membranes to help elicit fusion because of the amphipathic nature of their helices (Segrest et al, 1992;Sergel-Germano et al, 1994). Synthetic peptides representing part of the influenza virus HA heptad repeat region have been shown to insert reversibly into phospholipid vesicles under endosomal pH conditions (Yu et al, 1994).…”

Section: Discussionmentioning

confidence: 99%

Roles in Cell-to-Cell Fusion of Two Conserved Hydrophobic Regions in the Murine Coronavirus Spike Protein

1998

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The spike (S) protein of coronavirus mouse hepatitis virus (MHV), mediates attachment and fusion during viral entry and cell-to-cell fusion later in infection. By analogy with other viral proteins that induce cell fusion the MHV S protein would be expected to have a hydrophobic stretch of amino acids that serves as a fusion peptide. Sequence analysis suggests that the S protein falls within the group of fusion proteins having internal rather than N-terminal fusion peptides. Based on the features of known viral fusion peptides, we identified two regions (PEP1 and PEP2) of MHV-A59 S2 as possible fusion peptides. Site-directed mutagenesis and an in viro cell-to-cell fusion assay were used to evaluate the roles of PEP1 and PEP2, as well as a third previously identified putative fusion domain (PEP3) in membrane fusion. Substitution of bulky hydrophobic residues with charged residues within PEP1 affects the fusion activity of the S protein without affecting processing and surface expression. Similar substitutions within PEP2 result in a fusion-negative phenotype; however, these mutant S proteins also exhibit defects in protein processing and surface expression which likely explain the loss of the ability to induce fusion. Thus PEP1 remains a candidate fusion peptide, while PEP2 may play a significant role in the overall structure or oligomerization of the S protein. PEP3 is an unlikely putative fusion peptide since it is not conserved among coronaviruses and nonconservative amino acid substitutions in PEP3 have minimal effects on cell-to-cell fusion.

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“…The HN glycoprotein is involved in attachment and release of virus and the F glycoprotein mediates fusion of the viral envelope with cellular membranes [6]. Mutational analysis has shown that the fusion peptide and the adjacent heptad repeat region play a role in the fusion activity of F protein [7]. This protein is synthesized as a precursor, F 0 , which must be proteolytically cleaved to F 1 and F 2 to activate fusion [6,8].…”

Section: Introductionmentioning

confidence: 99%

Role of fusion protein cleavage site in the virulence of Newcastle disease virus

Panda

Huang

Elankumaran

et al. 2004

Microbial Pathogenesis

273

221

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Newcastle disease virus (NDV) causes a highly contagious and economically important disease in poultry. Viral determinants of NDV virulence are not completely understood. The amino acid sequence at the protease cleavage site of the fusion (F) protein has been postulated as a major determinant of NDV virulence. In this study, we have examined the role of F protein cleavage site sequence in NDV virulence using reverse genetics technology. The sequence G-R-Q-G-R present at the cleavage site of the F protein of avirulent strain LaSota was mutated to R-R-Q-K-R, which is present in the F cleavage site of neurovirulent strain Beaudette C (BC). The resultant mutated LaSota V.F. virus did not require exogenous protease for infectivity in cell culture, indicating that the F protein was cleaved by intracellular proteases. The virulence of the mutant and parental viruses was evaluated in vivo by intracerebral pathogenicity index (ICPI) and intravenous pathogenicity index (IVPI) tests in chickens. Our results showed that the modification of the F protein cleavage site resulted in a dramatic increase in virulence from an ICPI value of 0.00 for LaSota to a value of 1.12 for LaSota V.F. However, the ICPI value of LaSota V.F. was lower than that of BC, which had a value of 1.58. Interestingly, the IVPI tests showed values of 0.00 for both LaSota and LaSota V.F. viruses, compared to the IVPI value of 1.45 of BC. In vitro characteristics of the viruses were also studied. Our results demonstrate that the efficiency of cleavage of the F protein plays an important role if the NDV is delivered directly into the brains of chicks, but there could be other viral factors that probably affect peripheral replication, viremia, or entry into the central nervous system.

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Mutations in the fusion peptide and heptad repeat regions of the Newcastle disease virus fusion protein block fusion

Cited by 89 publications

References 28 publications

Spring-Loaded Heptad Repeat Residues Regulate the Expression and Activation of Paramyxovirus Fusion Protein

Spring-Loaded Heptad Repeat Residues Regulate the Expression and Activation of Paramyxovirus Fusion Protein

Roles in Cell-to-Cell Fusion of Two Conserved Hydrophobic Regions in the Murine Coronavirus Spike Protein

Role of fusion protein cleavage site in the virulence of Newcastle disease virus

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