Conformational mapping of the N-terminal peptide of HIV-1 gp41 in membrane environments using 13C-enhanced Fourier transform infrared spectroscopy

Gordon, Larry M.; Mobley, Patrick W.; Pilpa, Rosemarie; Sherman, Mark A.; Waring, Alan J.

doi:10.1016/s0005-2736(01)00443-6

Cited by 76 publications

(152 citation statements)

References 102 publications

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“…Infrared structural investigations of membrane-associated HFP have generally been consistent with predominant antiparallel β-sheet conformation and were based on analysis of the wavenumbers of the amide I transition. 17,51,81,82 One infrared study proposed that there was β-hairpin structure in the Ala-1 to Gly-16 region but the present work did not support this model because there were no residues in this region with non-β-strand 13 C shifts. 51 To our knowledge, the complete 13 C shift assignment of the present study is the first definitive evidence for a fully extended conformation.…”

Section: Discussioncontrasting

confidence: 85%

Solid-State NMR Spectroscopy of Human Immunodeficiency Virus Fusion Peptides Associated with Host-Cell-Like Membranes: 2D Correlation Spectra and Distance Measurements Support a Fully Extended Conformation and Models for Specific Antiparallel Strand Registries

2008

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The human immunodeficiency virus (HIV) is "enveloped" by a membrane, and infection of a host cell begins with fusion between viral and target cell membranes. Fusion is catalyzed by the HIV gp41 protein which contains a functionally critical ~20-residue apolar "fusion peptide" (HFP) that associates with target cell membranes. In this study, chemically synthesized HFPs were associated with host-cell-like membranes and had "scatter-uniform" labeling (SUL), that is, only one residue of each amino acid type was U-13 C, 15 N labeled. For the first sixteen HFP residues, an unambiguous 13 C chemical shift assignment was derived from 2D 13 C/ 13 C correlation spectra with short mixing times, and the shifts were consistent with continuous β-strand conformation. 13 C-13 C contacts between residues on adjacent strands were derived from correlation spectra with long mixing times and suggested close proximity of the following residues: Ala-6/Gly-10, Ala-6/Phe-11, and Ile-4/Gly-13. Specific antiparallel β-strand registries were further tested using a set of HFPs that were 13 CO-labeled at Ala-14 and 15 N-labeled at either Val-2, Gly-3, Ile-4, or Gly-5. The solid-state NMR data were fit with 50-60% population of antiparallel HFP with either Ala-14/Gly-3 or Ala-14/Ile-4 registries and 40-50% population of structures not specified by the NMR experiments. The first two registries correlated with intermolecular hydrogen bonding of 15-16 apolar N-terminal residues and this hydrogen-bonding pattern would be consistent with a predominant location of these residues in the hydrophobic membrane interior. To our knowledge, these results provide the first residue-specific structural models for membrane-associated HFP in its β-strand conformation.Correspondence to: David P. Weliky, weliky@chemistry.msu.edu. Supporting Information Available: 2D PDSD spectra of HFP-F at 10 and 500 ms exchange times; 1D slices for the 2D PDSD spectra of HFP-C and HFP-D; REDOR spectra at 24 and 32 ms dephasing times for HFP-I and HFP-K samples prepared with 25 µM initial HFP concentration; derivation of (ΔS/S 0 ) cor from (ΔS/S 0 ) exp and σ cor from σ exp ; comparison of (ΔS/S 0 ) exp with (ΔS/S 0 ) cor for the HFP-H, I, J, and K samples; model geometries for the HFP-H, HFP-J, and HFP-K samples; and (ΔS/S 0 ) sim for all models for the HFP-H, HFP-I, HFP-J, and HFP-K samples. This material is available free of charge via the Internet at

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Section: Discussioncontrasting

confidence: 85%

Solid-State NMR Spectroscopy of Human Immunodeficiency Virus Fusion Peptides Associated with Host-Cell-Like Membranes: 2D Correlation Spectra and Distance Measurements Support a Fully Extended Conformation and Models for Specific Antiparallel Strand Registries

2008

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“…This result correlated with previous studies which supported the following structural features: (1) β strand HFP was fully extended between A1 and G16; (2) β strand HFP formed hydrogen bonded oligomers or aggregates; and (3) a significant fraction of the oligomers have an antiparallel arrangement with adjacent strand crossing between F8 and L9 (25,(27)(28)(29)35,37,85,86). Some of these studies also supported conformational disorder at A21 (27,28).…”

Section: Insertion Modelssupporting

confidence: 89%

Solid-State Nuclear Magnetic Resonance Measurements of HIV Fusion Peptide to Lipid Distances Reveal the Intimate Contact of β Strand Peptide with Membranes and the Proximity of the Ala-14−Gly-16 Region with Lipid Headgroups

2007

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Human immunodeficiency virus (HIV) infection begins with fusion between viral and host cell membranes and is catalyzed by the HIV gp41 fusion protein. The ~20 N-terminal apolar residues of gp41 are called the HIV fusion peptide (HFP), interact with the host cell membrane, and play a key role in fusion. In this study, the membrane location of peptides which contained the HFP sequence AVGIGALFLGFLGAAGSTMGARS was probed in samples containing either only phospholipids or phospholipids and cholesterol. Four HFPs were examined which each contained 13 CO labeling at three sequential residues between G5 and G16. The 13 CO chemical shifts indicated that HFP had predominant β strand conformation over the labeled residues in the samples. The internuclear distances between the HFP 13 COs and the lipid 31 Ps were measured using solid-state nuclear magnetic resonance rotational-echo double resonance experiments. The closest 13 CO-31 P distances of 5-6 Å were observed for HFP labeled between A14 and G16 and correlated with intimate association of β strand HFP and membranes. These results were confirmed with measurements using HFPs singly 13 CO labeled at A6 or A14. To our knowledge, these data are the first measurements of distances between HIV fusion peptide nuclei and lipid P and qualitative models of membrane location of oligomeric β strand HFP are presented which are consistent with the experimental data. Observation of intimate contact between β strand HFP and membranes provides rationale for further investigation of the relationship between structure and fusion activity for this conformation. KeywordsHIV; fusion peptide; membrane; carbonyl; phosphorus; REDOR; NMR The infection of enveloped viruses such as human immunodeficiency virus (HIV) begins with fusion between the viral and host cell membranes (1-4). Fusion may be catalyzed by fusion proteins and several models of fusion protein catalysis have been proposed (2,(5)(6)(7). For HIV, fusion is catalyzed by a "gp160" glycoprotein complex which is incorporated in the virus membrane and is composed of two non-covalently associated subunits "gp120" and "gp41". The gp120 subunit lies outside the virus and binds to receptors in the target cell membrane and the gp41 subunit contains a region inside HIV as well as a single-pass transmembrane domain *To whom correspondence should be addressed. Telephone: 517-355-9715. Fax: 517-353-1793. Email: weliky@chemistry.msu.edu. † This work was supported by NIH award AI47153 to D. P. W. NIH Public Access Author ManuscriptBiochemistry. Author manuscript; available in PMC 2009 January 27. NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author Manuscript (8,9). The ~170-residue ectodomain of gp41 lies outside HIV and is subdivided into a more C-terminal "soluble ectodomain" and a ~20-residue N-terminal fusion peptide (HFP) which is apolar and fairly conserved. The HFP is believed to interact with the target cell membrane after gp120 binds to cellular receptors and fusion is greatly disrupted by mutation or deletion of the H...

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“…It has therefore been hypothesized that during viral/target cell fusion, at least three HFPs insert into the target cell membrane with their C-termini in close proximity. There is some antibodybased evidence for this hypothesis (21).A variety of experimental methods have shown that membrane-associated HFP can assume either helical or nonhelical structure (5,7,(22)(23)(24)(25)(26)(27)(28)(29)(30)(31)(32). Models for the helical structure have been developed based on nuclear magnetic resonance (NMR), electron spin resonance (ESR), infrared (IR), and circular dichroism (CD) data, as well as computer simulations (33)(34)(35)(36)(37)(38)(39).…”

mentioning

confidence: 99%

Conformational Flexibility and Strand Arrangements of the Membrane-Associated HIV Fusion Peptide Trimer Probed by Solid-State NMR Spectroscopy

et al. 2006

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The human immunodeficiency virus (HIV) fusion peptide (HFP) is the N-terminal apolar region of the HIV gp41 fusion protein and interacts with target cell membranes and promotes membrane fusion. The free peptide catalyzes vesicle fusion at least to the lipid mixing stage and serves as a useful model fusion system. For gp41 constructs which lack the HFP, high-resolution structures show trimeric protein and suggest that at least three HFPs interact with the membrane with their C-termini in close proximity. In addition, previous studies have demonstrated that HFPs which are cross-linked at their C-termini to form trimers (HFPtr) catalyze fusion at a rate which is 15−40 times greater than noncross-linked HFP. In the present study, the structure of membrane-associated HFPtr was probed with solid-state nuclear magnetic resonance (NMR) methods. Chemical shift and intramolecular 13 CO-15 N distance measurements show that the conformation of the Leu-7 to Phe-11 region of HFPtr has predominant helical conformation in membranes without cholesterol and β strand conformation in membranes containing ∼30 mol% cholesterol. Interstrand 13 CO-13 CO and 13 CO-15 N distance measurements were not consistent with an in-register parallel strand arrangement but were consistent with either: (1) parallel arrangement with adjacent strands tworesidues out-of-register; or (2) antiparallel arrangement with adjacent strand crossing between Phe-8 and Leu-9. Arrangement (1) could support the rapid fusion rate of HFPtr because of placement of the apolar N-terminal regions of all strands on the same side of the oligomer while arrangement (2) could support the assembly of multiple fusion protein trimers. KeywordsHIV; fusion peptide; cholesterol; membranes; NMR; trimer Enveloped viruses such as human immunodeficiency virus (HIV) are surrounded by a membrane and infect cells through fusion between the viral membrane and the target cell membrane (1,2). This process is mediated by envelope proteins which traverse the viral membrane (3). For HIV, the gp41 envelope protein has a ∼170-residue ectodomain region † This work was supported by NIH award AI47153 to D. P. W. * To whom correspondence should be addressed. Telephone: 517−355−9715. Fax: 517−353−1793. Email: weliky@chemistry.msu.edu.. SUPPORTING INFORMATION AVAILABLE Equations are derived for determination of (ΔS/S 0 ) cor for REDOR and fpCTDQBU analyses. This material is available free of charge via the Internet at http://pubs.acs.org. NIH Public Access Author ManuscriptBiochemistry. Author manuscript; available in PMC 2008 October 20. NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author Manuscript which lies outside the viral membrane and contains a N-terminal ∼20-residue apolar "fusion peptide" (HFP). The HFP interacts with the target cell membrane and plays an essential role in membrane fusion (4). Peptides composed of the HFP sequence induce fusion between large unilamellar vesicles (LUVs) and between erythrocytes (5,6). There are similar mutation/fusion activity relationships...

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Conformational mapping of the N-terminal peptide of HIV-1 gp41 in membrane environments using 13C-enhanced Fourier transform infrared spectroscopy

Cited by 76 publications

References 102 publications

Solid-State NMR Spectroscopy of Human Immunodeficiency Virus Fusion Peptides Associated with Host-Cell-Like Membranes: 2D Correlation Spectra and Distance Measurements Support a Fully Extended Conformation and Models for Specific Antiparallel Strand Registries

Solid-State NMR Spectroscopy of Human Immunodeficiency Virus Fusion Peptides Associated with Host-Cell-Like Membranes: 2D Correlation Spectra and Distance Measurements Support a Fully Extended Conformation and Models for Specific Antiparallel Strand Registries

Solid-State Nuclear Magnetic Resonance Measurements of HIV Fusion Peptide to Lipid Distances Reveal the Intimate Contact of β Strand Peptide with Membranes and the Proximity of the Ala-14−Gly-16 Region with Lipid Headgroups

Conformational Flexibility and Strand Arrangements of the Membrane-Associated HIV Fusion Peptide Trimer Probed by Solid-State NMR Spectroscopy

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