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dc.contributor.authorSergel, Theresa A.
dc.contributor.authorCullen, Lori McGinnes
dc.contributor.authorMorrison, Trudy G.
dc.date2022-08-11T08:09:34.000
dc.date.accessioned2022-08-23T16:35:57Z
dc.date.available2022-08-23T16:35:57Z
dc.date.issued2001-08-03
dc.date.submitted2009-03-26
dc.identifier.citationJ Virol. 2001 Sep;75(17):7934-43.
dc.identifier.issn0022-538X (Print)
dc.identifier.pmid11483738
dc.identifier.urihttp://hdl.handle.net/20.500.14038/38676
dc.description.abstractParamyxovirus fusion proteins have two heptad repeat domains, HR1 and HR2, which have been implicated in the fusion activity of the protein. Peptides with sequences from these two domains form a six-stranded coiled coil, with the HR1 sequences forming a central trimer (K. A. Baker, R. E. Dutch, R. A. Lamb, and T. S. Jardetzky, Mol. Cell 3:309-319, 1999; X. Zhao, M. Singh, V. N. Malashkevich, and P. S. Kim, Proc. Natl. Acad. Sci. USA 97:14172-14177, 2000). We have extended our previous mutational analysis of the HR1 domain of the Newcastle disease virus fusion protein, focusing on the role of the amino acids forming the hydrophobic core of the trimer, amino acids in the "a" and "d" positions of the helix from amino acids 123 to 182. Both conservative and nonconservative point mutations were characterized for their effects on synthesis, stability, proteolytic cleavage, and surface expression. Mutant proteins expressed on the cell surface were characterized for fusion activity by measuring syncytium formation, content mixing, and lipid mixing. We found that all mutations in the "a" position interfered with proteolytic cleavage and surface expression of the protein, implicating the HR1 domain in the folding of the F protein. However, mutation of five of seven "d" position residues had little or no effect on surface expression but, with one exception at residue 175, did interfere to various extents with the fusion activity of the protein. One of these "d" mutations, at position 154, interfered with proteolytic cleavage, while the rest of the mutants were cleaved normally. That most "d" position residues do affect fusion activity argues that a stable HR1 trimer is required for formation of the six-stranded coiled coil and, therefore, optimal fusion activity. That most of the "d" position mutations do not block folding suggests that formation of the core trimer may not be required for folding of the prefusion form of the protein. We also found that mutations within the fusion peptide, at residue 128, can interfere with folding of the protein, implicating this region in folding of the molecule. No characterized mutation enhanced fusion.
dc.language.isoen_US
dc.relation<a href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=pubmed&cmd=Retrieve&list_uids=11483738&dopt=Abstract">Link to Article in PubMed</a>
dc.subjectAmino Acid Sequence
dc.subjectAnimals
dc.subjectCOS Cells
dc.subjectCentrifugation, Density Gradient
dc.subjectCercopithecus aethiops
dc.subjectFlow Cytometry
dc.subjectFluorescent Antibody Technique
dc.subjectMolecular Sequence Data
dc.subject*Mutation
dc.subjectNewcastle disease virus
dc.subjectPeptides
dc.subject*Protein Folding
dc.subjectViral Fusion Proteins
dc.subjectLife Sciences
dc.subjectMedicine and Health Sciences
dc.titleMutations in the fusion peptide and adjacent heptad repeat inhibit folding or activity of the Newcastle disease virus fusion protein
dc.typeJournal Article
dc.source.journaltitleJournal of virology
dc.source.volume75
dc.source.issue17
dc.identifier.legacyfulltexthttps://escholarship.umassmed.edu/cgi/viewcontent.cgi?article=2531&amp;context=oapubs&amp;unstamped=1
dc.identifier.legacycoverpagehttps://escholarship.umassmed.edu/oapubs/1532
dc.identifier.contextkey798508
refterms.dateFOA2022-08-23T16:35:57Z
html.description.abstract<p>Paramyxovirus fusion proteins have two heptad repeat domains, HR1 and HR2, which have been implicated in the fusion activity of the protein. Peptides with sequences from these two domains form a six-stranded coiled coil, with the HR1 sequences forming a central trimer (K. A. Baker, R. E. Dutch, R. A. Lamb, and T. S. Jardetzky, Mol. Cell 3:309-319, 1999; X. Zhao, M. Singh, V. N. Malashkevich, and P. S. Kim, Proc. Natl. Acad. Sci. USA 97:14172-14177, 2000). We have extended our previous mutational analysis of the HR1 domain of the Newcastle disease virus fusion protein, focusing on the role of the amino acids forming the hydrophobic core of the trimer, amino acids in the "a" and "d" positions of the helix from amino acids 123 to 182. Both conservative and nonconservative point mutations were characterized for their effects on synthesis, stability, proteolytic cleavage, and surface expression. Mutant proteins expressed on the cell surface were characterized for fusion activity by measuring syncytium formation, content mixing, and lipid mixing. We found that all mutations in the "a" position interfered with proteolytic cleavage and surface expression of the protein, implicating the HR1 domain in the folding of the F protein. However, mutation of five of seven "d" position residues had little or no effect on surface expression but, with one exception at residue 175, did interfere to various extents with the fusion activity of the protein. One of these "d" mutations, at position 154, interfered with proteolytic cleavage, while the rest of the mutants were cleaved normally. That most "d" position residues do affect fusion activity argues that a stable HR1 trimer is required for formation of the six-stranded coiled coil and, therefore, optimal fusion activity. That most of the "d" position mutations do not block folding suggests that formation of the core trimer may not be required for folding of the prefusion form of the protein. We also found that mutations within the fusion peptide, at residue 128, can interfere with folding of the protein, implicating this region in folding of the molecule. No characterized mutation enhanced fusion.</p>
dc.identifier.submissionpathoapubs/1532
dc.contributor.departmentDepartment of Molecular Genetics and Microbiology
dc.source.pages7934-43


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