Synaptotagmin 1 and Ca2+ drive trans SNARE zippering.
Synaptotagmin 1 (Syt1) is a major Ca(2+)-sensor that evokes neurotransmitter release. Here we used site-specific fluorescence resonance energy transfer (FRET) assay to investigate the effects of Syt1 on SNAREpin assembly. C2AB, a soluble version of Syt1, had virtually no stimulatory effect on the rate of the FRET at N-terminus of SNARE complex both with and without Ca(2+), indicating C2AB does not interfere with the initial nucleation of SNARE assembly. However, C2AB-Ca(2+) accelerated the FRET rate significantly at membrane proximal region, indicating C2AB-Ca(2+) promotes the transition from a partially assembled SNARE complex to the fusion-competent SNAREpin. Similar enhancement was also observed at the end of the transmembrane domain of SNARE proteins. The stimulatory effect disappeared if there was no membrane or only neutral membrane present.
1] Department of Biochemistry, Biophysics, and Molecular Biology, Iowa State University, Ames, Iowa 50011, United States [2].
Ying
Y
Lai
Department of Biochemistry, Biophysics, and Molecular Biology, Iowa State University, Ames, Iowa 50011, United States.
Xiaochu
X
Lou
Cornell University, Ithaca, New York 14853, United States.
Chuqi
C
Wang
Department of the Electronics and Information Engineering, Huazhong University of Science and Technology, Wuhan, Hubei 430074, China.
Tian
T
Xia
Department of Cellular and Molecular Biology, The Scripps Research Institute, La Jolla, California 92037, United States.
Jiansong
J
Tong
D002118:Calcium / Q000378:metabolism*; D031541:Fluorescence Resonance Energy Transfer; D006801:Humans; D008027:Light; D008051:Lipid Bilayers / Q000737:chemistry / Q000378:metabolism; D008958:Models, Molecular; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D050600:SNARE Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012542:Scattering, Radiation; D050863:Synaptotagmin I / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
D008051:Lipid Bilayers; D011993:Recombinant Fusion Proteins; D050600:SNARE Proteins; D050863:Synaptotagmin I; D002118:Calcium