The location and nature of calcium-binding sites in salivary acidic proline-rich phosphoproteins.
The location of the calcium-binding sites in the human acidic proline-rich proteins, salivary proteins A and C, were determined by equilibrium dialysis of the tryptic peptides with buffers containing 45Ca. All the calcium-binding sites are located in the NH2-terminal tryptic peptide (TX peptide). The nature of the calcium binding sites in the TX peptide and native salivary proteins A and C, as well as dephosphorylated proteins were compared. Two types of sites can be distinguished in peptide TX. Type I sites have an apparent dissociation constant (K) of 38 microM and are responsible for the binding of 2.6 mol of Ca/mol of peptide. The corresponding figures for Type II sites are 780 microM and 5.3 mol of Ca/mol of peptide. In the native proteins, the amount of calcium bound at the type II sites decreases to 3.9 mol of Ca/mol of proteins A and C and K increases to 1100 microM. The amount of calcium bound at type I sites decreases to 1.5 mol/mol of protein A and 0.6 mol/mol of protein C, but there is no change in K. Dephosphorylation affects the calcium binding at both types of sites. The experiments indicate that the COOH-terminal parts of the native proteins affect the number and the nature of the protein calcium-binding sites. Proton and phosphorous NMR data demonstrate that beta-COOH in aspartic acid, as well as phosphoserine, are part of the calcium-binding sites. The difference in calcium binding to salivary proteins A and C may be due at least partially to differences in the environment of one or more aspartic acids.
D000469:Alkaline Phosphatase; D001665:Binding Sites; D002118:Calcium / Q000378:metabolism*; D002135:Calcium-Binding Proteins / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D010455:Peptides / Q000378:metabolism*; D010750:Phosphoproteins / Q000378:metabolism*; D011392:Proline; D055232:Proline-Rich Protein Domains; D011485:Protein Binding; D055273:Salivary Proline-Rich Proteins; D012471:Salivary Proteins and Peptides / Q000378:metabolism*
D002135:Calcium-Binding Proteins; C524536:PRH1 protein, human; D010455:Peptides; D010750:Phosphoproteins; D055273:Salivary Proline-Rich Proteins; D012471:Salivary Proteins and Peptides; D011392:Proline; D000469:Alkaline Phosphatase; D002118:Calcium