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Conformational flexibility and strand arrangements of the membrane-associated HIV fusion peptide trimer probed by solid-state NMR spectroscopy.

ABSTRACT: 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 that of non-cross-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 beta strand conformation in membranes containing approximately 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 two residues 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.

SUBMITTER: Zheng Z

PROVIDER: S-EPMC2570372 | biostudies-literature | 2006 Oct

REPOSITORIES: biostudies-literature

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Conformational flexibility and strand arrangements of the membrane-associated HIV fusion peptide trimer probed by solid-state NMR spectroscopy.

Zheng Zhaoxiong Z Yang Rong R Bodner Michele L ML Weliky David P DP

Biochemistry 20061001 43

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 pr ...[more]

PMID: 17059213

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Project description:Amyloid aggregates of a C-truncated Y145Stop mutant of human prion protein, huPrP23-144, associated with a heritable amyloid angiopathy, have previously been shown to contain a compact, relatively rigid, and beta-sheet-rich approximately 30-residue amyloid core near the C-terminus under physiologically relevant conditions. In contrast, the remaining huPrP23-144 residues display considerable conformational dynamics, as evidenced by the absence of corresponding signals in cross-polarization (CP)-based solid-state NMR (SSNMR) spectra under ambient conditions and their emergence in analogous spectra recorded at low temperature on frozen fibril samples. Here, we present the direct observation of residues comprising the flexible N-terminal domain of huPrP23-144 amyloid by using 2D J-coupling-based magic-angle spinning (MAS) SSNMR techniques. Chemical shifts for these residues indicate that the N-terminal domain is effectively an ensemble of protein chains with random-coil-like conformations. Interestingly, a detailed analysis of signal intensities in CP-based 3D SSNMR spectra suggests that non-negligible molecular motions may also be occurring on the NMR time scale within the relatively rigid core of huPrP23-144 amyloid. To further investigate this hypothesis, quantitative measurements of backbone dipolar order parameters and transverse spin relaxation rates were performed for the core residues. The observed order parameters indicate that, on the submicrosecond time scale, these residues are effectively rigid and experience only highly restricted and relatively uniform motions similar to those characteristic for well-structured regions of microcrystalline proteins. On the other hand, significant variations in magnitude of transverse spin relaxation rates were noted for residues present at different locations within the core region and correlated with observed differences in spectral intensities. While interpreted only qualitatively at the present time, the extent of the observed variations in transverse relaxation rates is consistent with the presence of relatively slow, microsecond-millisecond time scale chemical exchange type phenomena within the huPrP23-144 amyloid core.

Dataset Information

Conformational flexibility and strand arrangements of the membrane-associated HIV fusion peptide trimer probed by solid-state NMR spectroscopy.

Publications

Conformational flexibility and strand arrangements of the membrane-associated HIV fusion peptide trimer probed by solid-state NMR spectroscopy.

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