Isolation and characterization of peptides from the protein core of bovine gallbladder mucin.
Gallbladder mucin may promote cholesterol gallstone formation by accelerating cholesterol monohydrate crystal nucleation in supersaturated bile. In this study, peptides were isolated from the mucin protein core by protease digestion and molecular-sieve high-performance liquid chromatography. Tryptic peptides were purified by anion exchange or reverse-phase high-performance liquid chromatography, and amino acid compositions were determined. Tryptic peptides were (a) nonglycosylated, (b) selectively enriched in serine, glutamic acid plus glutamine, and glycine, and (c) depleted in threonine and proline compared with native gallbladder mucin. Bilirubin derivatized with Woodward's reagent K covalently bound to purified mucin. Tryptic digestion of the mucin-bilirubin complex yielded low-molecular-weight nonglycosylated peptides with covalently bound bilirubin. These data indicate that the mucin protein core contains at least two distinct domains. One domain is rich in threonine and proline and contains the majority of covalently bound carbohydrate. A second domain, possibly internally located, is nonglycosylated, enriched in serine, glutamic acid plus glutamine, and glycine, and binds hydrophobic ligands such as bilirubin and 1-anilino-8-naphthalene sulfonate. Hydrophobic domains on the mucin protein core may contribute to the pathogenesis of cholesterol cholelithiasis.
Section of Gastroenterology and Hepatology, Boston City Hospital, Massachusetts.
N H
NH
Afdhal
G D
GD
Offner
F E
FE
Murray
R F
RF
Troxler
B F
BF
Smith
D000596:Amino Acids / Q000032:analysis; D000817:Anilino Naphthalenesulfonates; D000818:Animals; D001663:Bilirubin; D002417:Cattle; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D005456:Fluorescent Dyes; D005704:Gallbladder / Q000378:metabolism*; D009077:Mucins / Q000032:analysis*; D010447:Peptide Hydrolases; D010455:Peptides / Q000302:isolation & purification*; D011485:Protein Binding; D011506:Proteins / Q000302:isolation & purification*; D012967:Sodium Dodecyl Sulfate; D013050:Spectrometry, Fluorescence
D000596:Amino Acids; D000817:Anilino Naphthalenesulfonates; D005456:Fluorescent Dyes; D009077:Mucins; D010455:Peptides; D011506:Proteins; D012967:Sodium Dodecyl Sulfate; C027132:1-anilino-8-naphthalenesulfonate; D010447:Peptide Hydrolases; D001663:Bilirubin