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)

• Published in: Biochimica et biophysica acta 2010-02, Vol.1798 (2), p.297-302
• Main Authors: Kathir, Karuppanan Muthusamy, Gao, Li, Rajalingam, Dakshinamurthy, Daily, Anna E., Brixey, Sherri, Liu, Huimin, Davis, Dan, Adams, Paul, Prudovsky, Igor, Kumar, Thallapuranam Krishnaswamy Suresh
• Format: Article
• Language: English
• Subjects: Copper - chemistry; Copper - metabolism; Fibroblast growth factor; Fibroblast Growth Factor 1 - chemistry; Fibroblast Growth Factor 1 - secretion; Humans; Lipid Bilayers - chemistry; Lipid Bilayers - metabolism; Lipid binding; Membrane Lipids - chemistry; Membrane Lipids - metabolism; Multiprotein Complexes - chemistry; Multiprotein Complexes - metabolism; Non-classical; Nuclear Magnetic Resonance, Biomolecular; Protein Structure, Quaternary - physiology; Protein Structure, Secondary - physiology; Secretion; Synaptotagmin; Synaptotagmin I - chemistry; Synaptotagmin I - metabolism
• ISSN: 0005-2736; 0006-3002; 1879-2642
• DOI: 10.1016/j.bbamem.2009.09.024

Human fibroblast growth factor (hFGF-1) is a ∼ 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 1H– 15N heteronuclear single quantum coherence experiments reveal that residues in β-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 β-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.