Skip to main content
  • ASM
    • Antimicrobial Agents and Chemotherapy
    • Applied and Environmental Microbiology
    • Clinical Microbiology Reviews
    • Clinical and Vaccine Immunology
    • EcoSal Plus
    • Infection and Immunity
    • Journal of Bacteriology
    • Journal of Clinical Microbiology
    • Journal of Microbiology & Biology Education
    • Journal of Virology
    • mBio
    • Microbiology and Molecular Biology Reviews
    • Microbiology Resource Announcements
    • Microbiology Spectrum
    • Molecular and Cellular Biology
    • mSphere
    • mSystems
  • Log in
  • My alerts
  • My Cart

Main menu

  • Home
  • Articles
    • Current Issue
    • Accepted Manuscripts
    • COVID-19 Special Collection
    • Minireviews
    • JVI Classic Spotlights
    • Archive
  • For Authors
    • Submit a Manuscript
    • Scope
    • Editorial Policy
    • Submission, Review, & Publication Processes
    • Organization and Format
    • Errata, Author Corrections, Retractions
    • Illustrations and Tables
    • Nomenclature
    • Abbreviations and Conventions
    • Publication Fees
    • Ethics Resources and Policies
  • About the Journal
    • About JVI
    • Editor in Chief
    • Editorial Board
    • For Reviewers
    • For the Media
    • For Librarians
    • For Advertisers
    • Alerts
    • RSS
    • FAQ
  • Subscribe
    • Members
    • Institutions
  • ASM
    • Antimicrobial Agents and Chemotherapy
    • Applied and Environmental Microbiology
    • Clinical Microbiology Reviews
    • Clinical and Vaccine Immunology
    • EcoSal Plus
    • Infection and Immunity
    • Journal of Bacteriology
    • Journal of Clinical Microbiology
    • Journal of Microbiology & Biology Education
    • Journal of Virology
    • mBio
    • Microbiology and Molecular Biology Reviews
    • Microbiology Resource Announcements
    • Microbiology Spectrum
    • Molecular and Cellular Biology
    • mSphere
    • mSystems

User menu

  • Log in
  • My alerts
  • My Cart

Search

  • Advanced search
Journal of Virology
publisher-logosite-logo

Advanced Search

  • Home
  • Articles
    • Current Issue
    • Accepted Manuscripts
    • COVID-19 Special Collection
    • Minireviews
    • JVI Classic Spotlights
    • Archive
  • For Authors
    • Submit a Manuscript
    • Scope
    • Editorial Policy
    • Submission, Review, & Publication Processes
    • Organization and Format
    • Errata, Author Corrections, Retractions
    • Illustrations and Tables
    • Nomenclature
    • Abbreviations and Conventions
    • Publication Fees
    • Ethics Resources and Policies
  • About the Journal
    • About JVI
    • Editor in Chief
    • Editorial Board
    • For Reviewers
    • For the Media
    • For Librarians
    • For Advertisers
    • Alerts
    • RSS
    • FAQ
  • Subscribe
    • Members
    • Institutions
Structure and Assembly

Membrane Structures of the Hemifusion-Inducing Fusion Peptide Mutant G1S and theFusion-Blocking Mutant G1V of Influenza Virus HemagglutininSuggest a Mechanism for Pore Opening in MembraneFusion

Yinling Li, Xing Han, Alex L. Lai, John H. Bushweller, David S. Cafiso, Lukas K. Tamm
Yinling Li
Department of Molecular Physiology and Biological Physics, University of Virginia, Charlottesville, Virginia
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Xing Han
Department of Molecular Physiology and Biological Physics, University of Virginia, Charlottesville, Virginia
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Alex L. Lai
Department of Molecular Physiology and Biological Physics, University of Virginia, Charlottesville, Virginia
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
John H. Bushweller
Department of Molecular Physiology and Biological Physics, University of Virginia, Charlottesville, Virginia
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
David S. Cafiso
Department of Molecular Physiology and Biological Physics, University of Virginia, Charlottesville, Virginia
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Lukas K. Tamm
Department of Molecular Physiology and Biological Physics, University of Virginia, Charlottesville, Virginia
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • For correspondence: lkt2e@virginia.edu
DOI: 10.1128/JVI.79.18.12065-12076.2005
  • Article
  • Figures & Data
  • Info & Metrics
  • PDF
Loading

Article Figures & Data

Figures

  • Tables
  • FIG. 1.
    • Open in new tab
    • Download powerpoint
    FIG. 1.

    Backbone 1H chemical shift differences indicating structural differences between mutant and wild-type fusion domains bound to DPC micelles at pH 5. A, Differences between G1S and wild-type fusion domains. B, Differences between G1V and wild-type fusion domains.

  • FIG. 2.
    • Open in new tab
    • Download powerpoint
    FIG. 2.

    Structures of G1S, G1V, and wild-type fusion domains in DPC micelles determined at pH 5 by 1H-NMR spectroscopy. (A) 20 conformers representing the structure of G1S are shown in blue. For comparison, the“ most typical” conformer of the wild-type fusion domain is superimposed in red on the family of G1S conformers. (B) Twenty conformers representing the structure of G1V are shown in blue. For comparison, the “most typical” conformer of the wild-type fusion domain is superimposed in red on the family of G1V conformers. (C) Ribbon representation of the closest-to-the-mean conformer of the G1S structure with side chains inserted. (D) Ribbon representation of the closest-to-the-mean conformer of the G1V structure with side chains inserted. (E) Ribbon representation of the closest-to-the-mean conformer of the wild-type structure with side chains inserted. (F) End-on views of the N-terminal helices of wild-type, G1S, and G1V fusion domains. The first, second, third, and fourth turns of the G1S helix are labeled. (G) GRASP (44)-generated electrostatic surface potential representations of the structures of wild-type, G1S, and G1V fusion domains at pH 5. Negative, positive, and neutral potentials are shown in red, blue, and white, respectively. Side, top, and bottom views are shown for each structure, and several residues are labeled for reference.

  • FIG. 3.
    • Open in new tab
    • Download powerpoint
    FIG. 3.

    Sections of NOESY spectra comparing NOEs that contribute to the definition of the different structures observed for the wild-type (WT), G1S, and G1V fusion domains in DPC micelles at pH 5. (A to C) NOE between Hα of Ile 10 and He3 of Trp 14 is present in the wild-type and G1S but absent in the G1V structure. (D to E) NOE between N-terminal Hα and HN of Phe 3 is present in G1S but very weak in the wild-type structure. (F to G) NOE between HN of Gly 13 and HN of Trp 14 is present in G1V but absent in the wild-type structure. All spectra were obtained under the same conditions and are plotted at the same contour level in each row. The thicker black lines are one-dimensional sections through the spectra at the positions of the indicated peaks.

  • FIG. 4.
    • Open in new tab
    • Download powerpoint
    FIG. 4.

    Depth of three critical spin-labeled residues (Phe 3, Glu 11, and Ile 18) of wild-type, G1S, and G1V fusion domains in POPC:POPG (4:1) bilayers at pH 5. The depths were determined by fitting EPR spectra at increasing microwave powers in the presence of O2, N2, and NiEDDA to power saturation curves and ratioing the half-saturation powers in the presence of the different spin-relaxation agents as described in Materials and Methods. Inset: Best-fit calculated distances from NMR structures (abscissa) to experimental EPR depth parameters (ordinate) for wild-type, G1S, G1V, and four spin-labeled lipids used for depth calibration. All data fit the theoretical fitting function Φ = Atanh[B(x− C] + D, where A, B, C, and D are constants as described by Frazier et al. (19).

  • FIG. 5.
    • Open in new tab
    • Download powerpoint
    FIG. 5.

    Wild-type(WT), G1S, and G1V fusion domain structures docked to POPC bilayers using the experimental depth data of Fig. 4. C termini are on the left and N termini are on the right. The polar lipid head groups and glycerol backbones are shown in orange, and the aliphatic side chains are shown in green in the molecular dynamic-simulated lipid bilayers.

  • FIG. 6.
    • Open in new tab
    • Download powerpoint
    FIG. 6.

    ATR-FTIR spectra of wild-type, G1S, and G1V peptides bound to supported bilayers of POPC:POPG (4:1) at increasing concentrations at pH 5. Spectra were recorded after successive injections, from bottom to top, of 10, 20, 40, 80, and 100 μg/ml peptide and subsequent flushing of the cell with deuterium oxide buffer. The band at 1,735 cm−1 arises from the lipid ester carbonyl groups, and the complex band between 1,600 and 1,700 cm−1 is the amide I′ band from the bound fusion domains.

  • FIG. 7.
    • Open in new tab
    • Download powerpoint
    FIG. 7.

    Boomerang mechanism of influenza virus hemagglutinin-mediated membrane fusion. (A) The pH-induced spring-loaded conformational change in the ectodomain (6, 7) (not shown) thrusts the three boomerang-shaped fusion domains into the target membrane, where 7.6 kcal/mol of free energy is gained for each inserted domain. (B) The ectodomains tilt relative to the viral membrane plane (23, 49) and the boomerangs retrieve the target membrane and bring it into close juxtaposition with the viral membrane. The extended C-terminal “leashes” of the HA2 subunit pack into the grooves of the newly extended triple coiled coils at the N terminus and thereby bring the truncated N and C termini of HA2 into close proximity (9). Lipid exchange between the proximal leaflets, but not between the distal leaflets of the bilayer, can occur at this stage, which sometimes is also referred to as the hemifused state (10, 36). The boomerang shape of the fusion domain is required for the transition from A to B. For simplicity, only one fusion and one TM domain are shown, although it is known that three fusion and TM domains from several trimers all participate in a single fusion pore (4, 5, 13, 40). (C) In this model the fusion and TM domains interact by virtue of the glycine edge of the fusion domain to open the fusion pore. We hypothesize that once the proximal monolayers are sufficiently perturbed, the fusion domains latch onto the TM domains and glide down the TM domains. They thereby perturb not only the proximal but also the distal monolayers and thus open a first conductive fusion pore (39, 44). This event requires a TM domain that contains at least 17 hydrophobic residues (3) and a smooth glycine edge on the fusion domain (43). The fusion pore eventually dilates and permits unrestricted lipid flow in both leaflets of the bilayer. Again, only one fusion and one TM domain are shown for simplicity.

Tables

  • Figures
  • TABLE 1.

    Chemical shifts and assignments of backbone and side chain protons of wild-type, G1S, and G1V fusion domains in DPC micelles at pH 5.0a

    Embedded Image
    • ↵ a Chemical shifts are given in parts per million.

  • TABLE 2.

    Structural statistics of the NMR structures of G1S and G1V

    ParameterG1SG1V
    Target function (Å)0.21 ± 0.030.13 ± 0.05
    Experimental NMR constraints
        NOE distance constraints145136
            Intraresidue5042
            Sequential4857
            Medium range4737
            Long range00
        Angle constraints (derived from HABAS)5356
            φ1919
            ψ1616
            χ11111
            χ2910
    NMR constraint violations
        NOE constraint violations
            Sum (Å)2.57 ± 0.181.76 ± 0.20
            Maximum (Å)0.09 ± 0.010.10 ± 0.01
        Angle constraint violations
            Sum (°)3.52 ± 0.761.28 ± 1.42
            Maximum (°)2.12 ± 0.360.79 ± 0.67
    AMBER energy (kcal/mol)−171.3 ± 7.6−241.6 ± 21.4
    Root mean squared deviation from the mean structure (Å)
        Backbone atoms of all residues 1-201.21 ± 0.491.76 ± 0.41
        All heavy atoms of all residues 1-201.60 ± 0.462.42 ± 0.44
        Backbone atoms of residues 2-180.63 ± 0.281.06 ± 0.36
        All heavy atoms of residues 2-181.14 ± 0.321.87 ± 0.49
    Ramachandran statistics analyzed using PROCHECK-NMR
        Residues in allowed regions (%)100.0100.0
        Residues in disallowed regions (%)0.00.0
PreviousNext
Back to top
Download PDF
Citation Tools
Membrane Structures of the Hemifusion-Inducing Fusion Peptide Mutant G1S and theFusion-Blocking Mutant G1V of Influenza Virus HemagglutininSuggest a Mechanism for Pore Opening in MembraneFusion
Yinling Li, Xing Han, Alex L. Lai, John H. Bushweller, David S. Cafiso, Lukas K. Tamm
Journal of Virology Aug 2005, 79 (18) 12065-12076; DOI: 10.1128/JVI.79.18.12065-12076.2005

Citation Manager Formats

  • BibTeX
  • Bookends
  • EasyBib
  • EndNote (tagged)
  • EndNote 8 (xml)
  • Medlars
  • Mendeley
  • Papers
  • RefWorks Tagged
  • Ref Manager
  • RIS
  • Zotero
Print

Alerts
Sign In to Email Alerts with your Email Address
Email

Thank you for sharing this Journal of Virology article.

NOTE: We request your email address only to inform the recipient that it was you who recommended this article, and that it is not junk mail. We do not retain these email addresses.

Enter multiple addresses on separate lines or separate them with commas.
Membrane Structures of the Hemifusion-Inducing Fusion Peptide Mutant G1S and theFusion-Blocking Mutant G1V of Influenza Virus HemagglutininSuggest a Mechanism for Pore Opening in MembraneFusion
(Your Name) has forwarded a page to you from Journal of Virology
(Your Name) thought you would be interested in this article in Journal of Virology.
CAPTCHA
This question is for testing whether or not you are a human visitor and to prevent automated spam submissions.
Share
Membrane Structures of the Hemifusion-Inducing Fusion Peptide Mutant G1S and theFusion-Blocking Mutant G1V of Influenza Virus HemagglutininSuggest a Mechanism for Pore Opening in MembraneFusion
Yinling Li, Xing Han, Alex L. Lai, John H. Bushweller, David S. Cafiso, Lukas K. Tamm
Journal of Virology Aug 2005, 79 (18) 12065-12076; DOI: 10.1128/JVI.79.18.12065-12076.2005
del.icio.us logo Digg logo Reddit logo Twitter logo CiteULike logo Facebook logo Google logo Mendeley logo
  • Top
  • Article
    • ABSTRACT
    • MATERIALS AND METHODS
    • RESULTS
    • DISCUSSION
    • ACKNOWLEDGMENTS
    • FOOTNOTES
    • REFERENCES
  • Figures & Data
  • Info & Metrics
  • PDF

KEYWORDS

Hemagglutinin Glycoproteins, Influenza Virus
influenza A virus
membrane fusion
Viral Fusion Proteins

Related Articles

Cited By...

About

  • About JVI
  • Editor in Chief
  • Editorial Board
  • Policies
  • For Reviewers
  • For the Media
  • For Librarians
  • For Advertisers
  • Alerts
  • RSS
  • FAQ
  • Permissions
  • Journal Announcements

Authors

  • ASM Author Center
  • Submit a Manuscript
  • Article Types
  • Ethics
  • Contact Us

Follow #Jvirology

@ASMicrobiology

       

 

JVI in collaboration with

American Society for Virology

ASM Journals

ASM journals are the most prominent publications in the field, delivering up-to-date and authoritative coverage of both basic and clinical microbiology.

About ASM | Contact Us | Press Room

 

ASM is a member of

Scientific Society Publisher Alliance

 

American Society for Microbiology
1752 N St. NW
Washington, DC 20036
Phone: (202) 737-3600

Copyright © 2021 American Society for Microbiology | Privacy Policy | Website feedback

Print ISSN: 0022-538X; Online ISSN: 1098-5514