title: 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).
abstract: 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.
authors.affiliation: Department of Chemistry and Biochemistry, University of Arkansas, Fayetteville, AR 72701, USA.
authors.forename: Karuppanan Muthusamy
authors.initials: KM
authors.lastname: Kathir
authors.forename: Li
authors.initials: L
authors.lastname: Gao
authors.forename: Dakshinamurthy
authors.initials: D
authors.lastname: Rajalingam
authors.forename: Anna E
authors.initials: AE
authors.lastname: Daily
authors.forename: Sherri
authors.initials: S
authors.lastname: Brixey
authors.forename: Huimin
authors.initials: H
authors.lastname: Liu
authors.forename: Dan
authors.initials: D
authors.lastname: Davis
authors.forename: Paul
authors.initials: P
authors.lastname: Adams
authors.forename: Igor
authors.initials: I
authors.lastname: Prudovsky
authors.forename: Thallapuranam Krishnaswamy Suresh
authors.initials: TK
authors.lastname: Kumar
mesh_terms: 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
chemical_list: D008051:Lipid Bilayers; D008563:Membrane Lipids; D046912:Multiprotein Complexes; C494310:SYT1 protein, human; D050863:Synaptotagmin I; D016220:Fibroblast Growth Factor 1; D003300:Copper