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The transmembrane domain peptide of vesicular stomatitis virus promotes both intermediate and pore formation during PEG-mediated vesicle fusion.

Abstract：

:We propose mechanisms by which the transmembrane domain of vesicular stomatitis virus (VSV-TMD) promotes both initiation of fusion and formation of a fusion pore. Time courses of polyethyleneglycol (PEG)-mediated fusion of 25 nm small unilamellar vesicles composed of dioleoylphosphatidylcholine, dioleoylphosphatidylethanolamine (DOPE), bovine brain sphingomyelin, and cholesterol (35:30:15:20 molar ratio) were recorded at pH 7.4 at five different temperatures (from 17°C to 37°C) and compared with time courses obtained with the same vesicles containing the fusion-active TMD of the G protein of VSV. Multiple time courses were fitted globally to a one-intermediate ensemble kinetic model to estimate the rate constants for conversion of the aggregated state to an intermediate hemifused state (k1, stalk, or I1) that rapidly transits to an unstable intermediate (I2 state) that converts to a final fusion pore state with a combined rate k3. The probabilities of lipid mixing, contents mixing, and contents leakage in the three states were also obtained from this analysis. The activation thermodynamics for each step were consistent with previously published models of lipid rearrangements during intermediate and pore formation. The influences of VSV-TMD, hexadecane, and VSV-TMD + hexadecane on the kinetics, activation thermodynamics, and membrane structure support the hypothesis that these two agents do not catalyze fusion by a common mechanism, except possibly at the lowest temperatures examined. VSV-TMD primarily catalyzed initial intermediate formation, although it substantially increased the probability of contents mixing in the intermediate state. Our results support the hypothesis that the catalytic influence of VSV-TMD on the initial-intermediate- and pore-forming steps of PEG-mediated fusion derives from its ability to impose a positive intrinsic curvature and thereby stress small unilamellar vesicle outer leaflets as well as the periphery of intermediate microstructures.

journal_name

Biophys J

journal_title

Biophysical journal

authors

Sengupta T,Chakraborty H,Lentz BR

doi

10.1016/j.bpj.2014.03.053

subject

Has Abstract

pub_date

2014-09-16 00:00:00

pages

1318-26

issue

eissn

0006-3495

issn

1542-0086

pii

S0006-3495(14)00794-2

journal_volume

107

pub_type

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The transmembrane domain peptide of vesicular stomatitis virus promotes both intermediate and pore formation during PEG-mediated vesicle fusion.