SOLID-PHASE SYNTHESIS AND CHARACTERIZATION OF HUMAN SALIVARY STATHERIN - A TYROSINE-RICH PHOSPHOPROTEIN INHIBITOR OF CALCIUM-PHOSPHATE PRECIPITATION

Human statherin, a low molecular weight (M(r) 5380 Da, 43 amino acid r esidues) acidic tyrosine-rich phosphoprotein secreted mainly by saliva ry glands, has been synthesized successfully for the first time follow ing standard solid-phase Fmoc chemistry. Synthesis of this phosphoprot ein was accomplished using preformed phosphoserine building blocks. Th e phosphorylated protein thus synthesized was analyzed and compared wi th the native molecule and was found to have identical characteristics in its entirety, as evidenced by various analytical methods including mass spectral analysis. Analysis of both the synthetic and native sta therin by circular dichroism spectroscopy showed an increase in helici ty upon the addition of an organic cosolvent, trifluoroethanol (50%, v ol/vol), indicating the presence of potentially amphipathic helical re gions. Circular dichroism studies and hydrophobic moment calculations on this synthetic phosphoprotein revealed that the molecule adopts an amphipathic helical conformation at the N terminus connected to a long poly-L-proline type II segment, which, in turn, is linked to an exten ded beta-strand. In correlation with previous studies, it appears that the strong binding affinity of statherin for hydroxyapatite can be at tributed primarily to the N-terminal sequence, which prefers to adopt a helical conformation and provides both electrostatic and hydrogen bo nding interactions, thereby inhibiting its mineralization. Production of this highly homogenous synthetic statherin by chemical means may ci rcumvent the prevailing obstacles encountered in conducting its tertia ry structural investigations under various physiological conditions.