| S100 proteins represent a group of small (about 100 amino acids) calcium-binding proteins that carry two calcium-binding motifs (EF-hands) in their primary sequence. Unlike another calcium-binding protein, calmodulin, S100 proteins exhibit tissue- and cell type-specific expression. S100 proteins have been frequently associated with a wide variety of neurological and neoplastic diseases. Although exact function of the S100 proteins is not clear, some of them are thought to play a role in regulating cytoskeletal dynamics, control of enzymatic activities, protein phosphorylation, lipid metabolism, control of cell survival and proliferation.; S100P is one of the least studied members of the S100 subfamily. Deregulation of S100P expression has been linked to the transformation of androgen-dependent prostate cancer into androgen-independent. Overexpression of S100P has been associated with immortalization of breast epithelial cells in vitro and breast tumor progression in vivo.; In order to clarify biological role of S100P, S100P cDNA was cloned into a bacterial expression vector, and the recombinant protein was overexpressed in E. coli. S100P calcium-binding properties were characterized using biophysical methods. Experiments with Y18F, Y88F and C85S variants of S100P led to the conclusion, that there are two calcium-binding sites in the S100P molecule. High-affinity site (with the affinity constant on the order of 107 M−1) is located in the C-terminal part of the molecule, while low-affinity site (with the affinity constant on the order of 104 M−1) is located in the N-terminus. The N-terminal site is also able to bind magnesium, and binding of magnesium increases Ca2+-binding affinity at the C-terminal site. Upon interaction with calcium, S100P undergoes conformational change, as can be judged from the CD and fluorescence spectroscopies, and analytical gel-filtration experiments. Based on these results a Ca2+/Mg 2+ conformational switch model was proposed.; Ca2+/Mg2+ switch hypothesis was confirmed using a model peptide melittin. S100P interacts with melittin in a calcium-dependent manner. In the absence of calcium, S100P binds melittin with low affinity, stoichiometry of 2 molecules of melittin bound per monomer of S100P, and the interaction probably involves negatively charged amino acid residues on the surface of S100P and positively charged residues in the C-terminus of melittin. In the presence of calcium, S100P binds melittin with almost 100-fold higher affinity, positive cooperativity, stoichiometry of one molecule of melittin bound per monomer of S100P, and the interaction involves hydrophobic surfaces that are exposed on S100P upon binding of the calcium ions. Ca2+-dependent interaction of S100P and melittin is accompanied by folding of the peptide into an amphipathic helix.; A search for the putative S100P-binding proteins by screening human prostate cDNA library using the yeast two-hybrid method, identified a novel member of the S100 subfamily, which we tentatively named S100Z. Isoelectric focusing experiments confirm that S100P and S100Z associate in vitro. S100Z is a 99 amino-acid protein, with high α-helix content, is capable of binding calcium and undergoes conformational change upon calcium binding. S100Z gene is located on chromosome 5. Using a variation of RT-PCR, S100Z mRNA was detected in several tissues, with the highest levels of expression found in leukocytes and spleen. Expression of the S100Z also appears to be aberrant in some tumors. |
