Interactions Defined between S100A13 and Annexin Peptides: Insight into Non-Classical Secrection

Abstract

S100A13 is a calcium binding chaperone protein, known to be involved in the non-classical export of signal peptide less proteins such as fibroblast growth factor (FGF-1) and interleukin-1α across the cell membrane. It has also been shown that the interaction of S100A13 with Annexin II, which exhibits an inducible flip-flop mechanism across the cell bilayer, helps the multiprotein release complex to traverse the membrane bilayer. The interaction of S100A13 and annexin II has been characterized using various biophysical techniques including multidimensional NMR spectroscopy. Results of the Isothermal titration calorimetry (ITC) experiments show that holo-S100A13 exhibits preferential binding to annexin II with high affinity in the micro molar rang compared to apo-S100A13. Equilibrium guanidine hydrochloride denaturation monitored by steady-state fluorescence and limited trypsin digestion analysis reveals holo-S100A13 is significantly stabilized upon binding to annexin II peptide compared to apo-S100A13. ANS (8-anilino-1-napthalene sulfonate) binding experiments indicates that the presence of annexin II peptide does not increase the solvent availability of hydrophobic residues in holo-S100A13. Availability of the solvent-exposed hydrophobic surface(s) in apoS10013 does not facilitate its interaction with the annexin II peptide, which is unique characteristic of S100A13. 1H-15N- HSQC NMR experiments reveal that the binding site of annexin II peptide on holo-S100A13 is modestly different from other S100/Annexin interactions. S100 proteins typically interact with more than one Annexin protein. In order to define the specificity of S100A13 for Annexin 2, the interaction of S100A13 with other Annexin peptides was characterized and data characterizing these interactions will be presented. Finally, defining the interaction between S100A13 and Annexin 2 peptide will give much needed insight into the non-classical release of the signal peptide-less protein Fibroblast Growth Factor-1