Studies of the membrane fusion activities of fusion peptide mutants of influenza virus hemagglutinin.

ABSTRACT Influenza virus hemagglutinin (HA)-mediated membrane fusion is initiated by a conformational change that releases a V-shaped hydrophobic fusion domain, the fusion peptide, into the lipid bilayer of the target membrane. The most N-terminal residue of this domain, a glycine, is highly conserved and is particularly critical for HA function; G1S and G1V mutant HAs cause hemifusion and abolish fusion, respectively. We have determined the atomic resolution structures of the G1S and G1V mutant fusion domains in membrane environments. G1S forms a V with a disrupted “glycine edge” on its N-terminal arm and G1V adopts a slightly tilted linear helical structure in membranes. Abolishment of the kink in G1V results in reduced hydrophobic penetration of the lipid bilayer and an increased propensity to formβ -structures at the membrane surface. These results underline the functional importance of the kink in the fusion peptide and suggest a structural role for the N-terminal glycine ridge in viral membrane fusion.

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Full-length trimeric influenza virus hemagglutinin II membrane fusion protein and shorter constructs lacking the fusion peptide or transmembrane domain: Hyperthermostability of the full-length protein and the soluble ectodomain and fusion peptide make significant contributions to fusion of membrane vesicles

Protein Expression and Purification ◽

10.1016/j.pep.2015.08.021 ◽

2016 ◽

Vol 117 ◽

pp. 6-16 ◽

Cited By ~ 6

Author(s):

Punsisi U. Ratnayake ◽

E.A. Prabodha Ekanayaka ◽

Sweta S. Komanduru ◽

David P. Weliky

Keyword(s):

Influenza Virus ◽

Fusion Protein ◽

Membrane Fusion ◽

Transmembrane Domain ◽

Membrane Vesicles ◽

Fusion Peptide ◽

Full Length ◽

Membrane Fusion Protein ◽

Influenza Virus Hemagglutinin

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The Stabilities of the Soluble Ectodomain and Fusion Peptide Hairpins of the Influenza Virus Hemagglutinin Subunit II Protein Are Positively Correlated with Membrane Fusion

Biochemistry ◽

10.1021/acs.biochem.8b00764 ◽

2018 ◽

Vol 57 (37) ◽

pp. 5480-5493 ◽

Cited By ~ 3

Author(s):

Ahinsa Ranaweera ◽

Punsisi U. Ratnayake ◽

David P. Weliky

Keyword(s):

Influenza Virus ◽

Membrane Fusion ◽

Fusion Peptide ◽

Influenza Virus Hemagglutinin

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Testing topological models for the membrane penetration of the fusion peptide of influenza virus hemagglutinin

FEBS Letters ◽

10.1016/0014-5793(89)81574-1 ◽

1989 ◽

Vol 257 (2) ◽

pp. 369-372 ◽

Cited By ~ 33

Author(s):

Josef Brunner

Keyword(s):

Influenza Virus ◽

Fusion Peptide ◽

Membrane Penetration ◽

Influenza Virus Hemagglutinin ◽

Topological Models

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New Insights into the Spring-Loaded Conformational Change of Influenza Virus Hemagglutinin

Journal of Virology ◽

10.1128/jvi.76.9.4456-4466.2002 ◽

2002 ◽

Vol 76 (9) ◽

pp. 4456-4466 ◽

Cited By ~ 42

Author(s):

Jennifer A. Gruenke ◽

R. Todd Armstrong ◽

William W. Newcomb ◽

Jay C. Brown ◽

Judith M. White

Keyword(s):

Influenza Virus ◽

Conformational Change ◽

Conformational Changes ◽

Limited Proteolysis ◽

Coiled Coil ◽

Fusion Peptide ◽

Wild Type ◽

Influenza Virus Hemagglutinin ◽

Target Membrane ◽

Mutant Forms

ABSTRACT Influenza virus hemagglutinin undergoes a conformational change in which a loop-to-helix “spring-loaded” conformational change forms a coiled coil that positions the fusion peptide for interaction with the target bilayer. Previous work has shown that two proline mutations designed to disrupt this change disrupt fusion but did not determine the basis for the fusion defect. In this work, we made six additional mutants with single proline substitutions in the region that undergoes the spring-loaded conformational change and two additional mutants with double proline substitutions in this region. All double mutants were fusion inactive. We analyzed one double mutant, F63P/F70P, as an example. We observed that F63P/F70P undergoes key low-pH-induced conformational changes and binds tightly to target membranes. However, limited proteolysis and electron microscopy observations showed that the mutant forms a coiled coil that is only ∼50% the length of the wild type, suggesting that it is splayed in its N-terminal half. This work further supports the hypothesis that the spring-loaded conformational change is necessary for fusion. Our data also indicate that the spring-loaded conformational change has another role beyond presenting the fusion peptide to the target membrane.

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