Open AccessA Central Small Amino Acid in the VAMP2 Transmembrane Domain Regulates the Fusion Pore in Exocytosis
Author(s) -
Benoît Hastoy,
Pier A. Scotti,
Alexandra Milochau,
Zahia Fezoua-Boubegtiten,
Jorge Rodas,
Rémi Mégret,
Bernard Desbat,
Michel S. Laguerre,
Sabine Castano,
David Perrais,
Patrik Rorsman,
Reïko Oda,
Jochen Lang
Publication year - 2017
Publication title -
scientific reports
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.24
H-Index - 213
ISSN - 2045-2322
DOI - 10.1038/s41598-017-03013-3
Subject(s) - exocytosis , biophysics , lipid bilayer fusion , transmembrane protein , vesicle fusion , amino acid , transmembrane domain , lipid bilayer , biology , membrane , chemistry , biochemistry , synaptic vesicle , vesicle , receptor
Exocytosis depends on cytosolic domains of SNARE proteins but the function of the transmembrane domains (TMDs) in membrane fusion remains controversial. The TMD of the SNARE protein synaptobrevin2/VAMP2 contains two highly conserved small amino acids, G 100 and C 103 , in its central portion. Substituting G 100 and/or C 103 with the β-branched amino acid valine impairs the structural flexibility of the TMD in terms of α-helix/β-sheet transitions in model membranes (measured by infrared reflection-absorption or evanescent wave spectroscopy) during increase in protein/lipid ratios, a parameter expected to be altered by recruitment of SNAREs at fusion sites. This structural change is accompanied by reduced membrane fluidity (measured by infrared ellipsometry). The G 100 V/C 103 V mutation nearly abolishes depolarization-evoked exocytosis (measured by membrane capacitance) and hormone secretion (measured biochemically). Single-vesicle optical (by TIRF microscopy) and biophysical measurements of ATP release indicate that G 100 V/C 103 V retards initial fusion-pore opening, hinders its expansion and leads to premature closure in most instances. We conclude that the TMD of VAMP2 plays a critical role in membrane fusion and that the structural mobility provided by the central small amino acids is crucial for exocytosis by influencing the molecular re-arrangements of the lipid membrane that are necessary for fusion pore opening and expansion.