Isolation of an Acidic Protein from Cholesterol Gallstones, which Inhibits the Precipitation of Calcium Carbonate in Vitro

In seeking to identify nucleating/antinucleating proteins in-volved in the pathogenesis of cholesterol gallstones, a major acidic protein was isolated from each of 13 samples of choles-terol gallstones. After the stones were extracted with methyl t-butyl ether to remove cholesterol, and methanol to remove bile salts and other lipids, they were demineralized with EDTA. The extracts were desalted with Sephadex-G25, and the proteins separated by PAGE. A protein was isolated, of molecular weight below 10 kD, which included firmly-bound diazo-positive yellow pigments and contained 24 % acidic, but only 7 % basic amino acid residues. The presence of N-acetyl glucosamine suggested that this was a glycoprotein. This pro-tein at concentrations as low as 2,g/ml, but neither human serum albumin nor its complex with bilirubin, inhibited cal-cium carbonate precipitation from a supersaturated solution in vitro. This protein could be precipitated from 0.15 M NaCI solution by the addition of 0.5 M calcium chloride. Considering that cholesterol gallstones contain calcium and pigment at their centers, and that small acidic proteins are important reg-ulators in other biomineralization systems, this protein seems likely to play a role in the pathogenesis of cholesterol gall-stones