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)

by Kathir, Karuppanan Muthusamy; Gao, Li; Dakshinamurthy, Rajalingam; Daily, Anna E.; Brixey, Sherri; Liu, Huimin; Davis, Dan J.; Adams, Paul D.; Prudovsky, Igor; Kumar, Thallapuranam Krishnaswamy Suresh

Human fibroblast growth factor (hFGF-1) is a similar to 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 Imiltiprotein complex consisting of hFGF-1, S100A13 (a calcium binding protein), and p40 synaptotagmin (Syt1). Copper (Cu2+) 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 Cu2+ and lipid interactions of the C2B domain of Syt1 using multidimensional NMR spectroscopy. The results highlight how Cu2+ appears to stabilize the protein bound to pS vesicles. Cu2+ and lipid binding interface mapped using 2D H-1-N-15 heteronuclear single quantum coherence experiments reveal that residues in beta-strand I contributes to the unique Cu2+ 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 Cu2+, 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 Cu2+-induced non-classical secretion of hFGF-1. (C) 2009 Elsevier B.V. All rights reserved.

Journal
Biochimica et Biophysica Acta-Biomembranes
Volume
1798
Issue
2
Year
2010
Start Page
297
ISBN/ISSN
0005-2736
PMID
19835837
DOI
10.1016/j.bbamem.2009.09.024