The long term objectives of this project are to investigate the structure and function of epithelial mucins and to investigate the contributions of these glycoproteins to disease states, particularly gallstones and peptic ulcer.
The specific aims of this application are focussed on the peptide core (apomucin) of gastric and gallbladder mucin in humans and experimental animals. We propose to: 1. Purify apomucin, obtain partial amino acid sequences and prepare apomucin-specific antibodies, 2. Use gallbladder and gastric RNA from several species to direct synthesis of apomucin in cell- free translating systems 3. Study lipid binding to apomucin and whole mucin, and determine the effects of bound lipid viscosity, barrier function, and cholesterol monohydrate nucleation. Our experiments will allow us to test the hypothesis that hydrophobic areas of apomucin are important for lipid binding which in turn influences functional parameter. Highly purified mucin from human stomachs and gallbladders, pig stomach and prairie dog gallbladder will be deglycosylated with trimethanesulfonic acid and endoglycosidases and purified by HPLC. Tryptic peptides of apomucin will be sequenced and antibodies to apomucin prepared in rabbits. Apomucin specific antibody will be used to identify translated apomucin in cell-free systems primed with gastric gallbladder and RNA. Purified apomucins from pig and human stomach will be evaluated for ability to bind lipids of various classes. These studies will provide for the first time the molecular weight, partial amino acid sequence and amino acid composition of intestinan and biliary apomucins and will allow comparison of apomucins from stomach and gallbladder in several species. Studies of lipid binding to apomucin will allow us to determine the site of binding, and the effects of bound lipid on functional parameters such as viscosity, ion diffusion and cholesterol monohydrate nucleation in model and native biles. Because mucin is thought to be involved in gallstone nucleation in man and experimental animals, these results should provide new insights into gallstone pathophysiology. These experiments should also provide a better understanding of how the mucus barrier protects the stomach from auto digestion.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK028195-10
Application #
3228661
Study Section
General Medicine A Subcommittee 2 (GMA)
Project Start
1980-07-01
Project End
1994-03-31
Budget Start
1990-04-01
Budget End
1991-03-31
Support Year
10
Fiscal Year
1990
Total Cost
Indirect Cost
Name
Boston University
Department
Type
DUNS #
City
Boston
State
MA
Country
United States
Zip Code
02118
Cao, X; Bansil, R; Bhaskar, K R et al. (1999) pH-dependent conformational change of gastric mucin leads to sol-gel transition. Biophys J 76:1250-8
Turner, B S; Bhaskar, K R; Hadzopoulou-Cladaras, M et al. (1999) Cysteine-rich regions of pig gastric mucin contain von willebrand factor and cystine knot domains at the carboxyl terminal(1). Biochim Biophys Acta 1447:77-92
Bhaskar, K R; Turner, B S; Grubman, S A et al. (1998) Dysregulation of proteoglycan production by intrahepatic biliary epithelial cells bearing defective (delta-f508) cystic fibrosis transmembrane conductance regulator. Hepatology 27:7-14
Grubel, P; Bhaskar, K R; Cave, D R et al. (1997) Interaction of an aluminum-magnesium containing antacid and gastric mucus: possible contribution to the cytoprotective function of antacids. Aliment Pharmacol Ther 11:139-45
Turner, B S; Bhaskar, K R; Hadzopoulou-Cladaras, M et al. (1995) Isolation and characterization of cDNA clones encoding pig gastric mucin. Biochem J 308 ( Pt 1):89-96
Afdhal, N H; Gong, D; Niu, N et al. (1993) Cholesterol cholelithiasis in the prairie dog: role of mucin and nonmucin glycoproteins. Hepatology 17:693-700
Lamont, J T; Carey, M C (1992) Cholesterol gallstone formation. 2. Pathobiology and pathomechanics. Prog Liver Dis 10:165-91
Carey, M C; Lamont, J T (1992) Cholesterol gallstone formation. 1. Physical-chemistry of bile and biliary lipid secretion. Prog Liver Dis 10:139-63
Bhaskar, K R; Gong, D H; Bansil, R et al. (1991) Profound increase in viscosity and aggregation of pig gastric mucin at low pH. Am J Physiol 261:G827-32
O'Leary, D P; Murray, F E; Turner, B S et al. (1991) Bile salts stimulate glycoprotein release by guinea pig gallbladder in vitro. Hepatology 13:957-61

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