Ions binding to S100 proteins. I. Calcium- and zinc-binding properties of bovine brain S100 alpha alpha, S100a (alpha beta), and S100b (beta beta) protein: Zn2+ regulates Ca2+ binding on S100b protein.
Flow dialysis measurements of calcium binding to bovine brain S100 alpha alpha, S100a (alpha beta), and S100b (beta beta) proteins in 20 mM Tris-HCl buffer at pH 7.5 and 8.3 revealed that S100 proteins bind specifically 4 Ca2+ eq/mol of protein dimer. The specific calcium-binding sites had, therefore, been assigned to typical amino acid sequences on the alpha and beta subunit. The protein affinity for calcium is much lower in the presence of magnesium and potassium. Potassium strongly antagonizes calcium binding on two calcium-binding sites responsible for most of the Ca2+-induced conformational changes on S100 proteins (probably site II alpha and site II beta). Zinc-binding studies in the absence of divalent cations revealed eight zinc-binding sites/mol of S100b protein dimer that we assumed to correspond to 4 zinc-binding sites/beta subunit. Zinc binding to S100b studied with UV spectroscopy methods showed that the occupation of the four higher affinity sites and the four lower affinity sites on the protein dimer were responsible for different conformational changes in S100b structure. Zinc binding on the higher affinity sites regulates calcium binding to S100b by increasing the protein affinity for calcium and decreasing the antagonistic effect of potassium on calcium binding. Zinc-binding studies on S100a and S100 alpha alpha protein showed that the Trp-containing S100 proteins bind zinc more weakly than S100b protein. Calcium-binding studies on zinc-bound S100a proved that calcium- and zinc-binding sites were distinct although there was no increase in zinc-bound S100a affinity for calcium, as in S100b protein. Finally we provide evidence that discrepancies between previously published results on the optical properties of S100b protein probably result from oxidation of the sulfhydryl groups in the protein.
D000818:Animals; D015415:Biomarkers; D001923:Brain Chemistry; D002118:Calcium / Q000378:metabolism*; D002417:Cattle; D006863:Hydrogen-Ion Concentration; D046911:Macromolecular Substances; D009414:Nerve Growth Factors; D011189:Potassium Chloride / Q000494:pharmacology; D064568:S100 Calcium Binding Protein beta Subunit; D009418:S100 Proteins / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D015032:Zinc / Q000378:metabolism*
D015415:Biomarkers; D046911:Macromolecular Substances; D009414:Nerve Growth Factors; C030564:S-100 calcium-binding protein alpha subunit; D064568:S100 Calcium Binding Protein beta Subunit; D009418:S100 Proteins; D011189:Potassium Chloride; D015032:Zinc; D002118:Calcium