NMR characterization of copper and lipid interactions of the C2B domain of synaptotagmin I-relevance to the non-classical secretion of the human acidic fibroblast growth factor (hFGF-1).
Human fibroblast growth factor (hFGF-1) is a approximately 17 kDa heparin binding cytokine. It lacks the conventional hydrophobic N-terminal signal sequence and is secreted through non-classical secretion routes. Under stress, hFGF-1 is released as a multiprotein complex consisting of hFGF-1, S100A13 (a calcium binding protein), and p40 synaptotagmin (Syt1). Copper (Cu(2+)) is shown to be required for the formation of the multiprotein hFGF-1 release complex (Landriscina et al. ,2001; Di Serio et al., 2008). Syt1, containing the lipid binding C2B domain, is believed to play an important role in the eventual export of the hFGF-1 across the lipid bilayer. In this study, we characterize Cu(2+) and lipid interactions of the C2B domain of Syt1 using multidimensional NMR spectroscopy. The results highlight how Cu(2+) appears to stabilize the protein bound to pS vesicles. Cu(2+) and lipid binding interface mapped using 2D (1)H-(15)N heteronuclear single quantum coherence experiments reveal that residues in beta-strand I contributes to the unique Cu(2+) binding site in the C2B domain. In the absence of metal ions, residues located in Loop II and beta-strand IV contribute to binding to unilamelar pS vesicles. In the presence of Cu(2+), additional residues located in Loops I and III appear to stabilize the protein-lipid interactions. The results of this study provide valuable information towards understanding the molecular mechanism of the Cu(2+)-induced non-classical secretion of hFGF-1.
Kathir
Karuppanan Muthusamy
KM
Department of Chemistry and Biochemistry, University of Arkansas, Fayetteville, AR 72701, USA.
Gao
Li
L
Rajalingam
Dakshinamurthy
D
Daily
Anna E
AE
Brixey
Sherri
S
Liu
Huimin
H
Davis
Dan
D
Adams
Paul
P
Prudovsky
Igor
I
Kumar
Thallapuranam Krishnaswamy Suresh
TK
D003300:Copper / Q000737:chemistry* / Q000378:metabolism; D016220:Fibroblast Growth Factor 1 / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D008051:Lipid Bilayers / Q000737:chemistry* / Q000378:metabolism; D008563:Membrane Lipids / Q000737:chemistry* / Q000378:metabolism; D046912:Multiprotein Complexes / Q000737:chemistry* / Q000378:metabolism; D019906:Nuclear Magnetic Resonance, Biomolecular; D020836:Protein Structure, Quaternary / Q000502:physiology; D017433:Protein Structure, Secondary / Q000502:physiology; D050863:Synaptotagmin I / Q000737:chemistry* / Q000378:metabolism
D008051:Lipid Bilayers; D008563:Membrane Lipids; D046912:Multiprotein Complexes; C494310:SYT1 protein, human; D050863:Synaptotagmin I; D016220:Fibroblast Growth Factor 1; D003300:Copper