Isolation and characterization of the native glycoprotein from pig small-intestinal mucus.
Glycoprotein from pig small-intestinal mucus was isolated free of non-covalently bound protein and nucleic acid with a yield of over 60%. No non-covalently bound protein could be detected by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis or by equilibrium centrifugation in a density gradient of CsCl with 4 M-guanidinium chloride. The intrinsic viscosity and reduced viscosity of the glycoprotein preparations rose with the removal of non-covalently bound protein and nucleic acid from the glycoprotein, evidence that non-covalently bound protein does not contribute to the rheological properties of the glycoprotein in the mucus. The pure glycoprotein, in contrast with impure preparations, gelled at the same concentration of glycoprotein as that present in the gel in vivo. The glycoprotein was a single component, as judged by gel filtration and analytical ultracentrifugation. The distribution of sedimentation coefficients was polydisperse but unimodal with an s025,w of 14.5S and a molecular weight of 1.72 X 10(6). The chemical composition of the glycoprotein was 77% carbohydrate and 21% protein, 52% of which was serine, threonine and proline. The glycoprotein had a strong negative charge and contained 3.1% and 18.3% by weight ester sulphate and sialic acid respectively. The molar proportion of N-acetylgalactosamine was nearly twice that of any of the other sugars present, the glycoprotein had A and H blood-group activity and the average maximum length of the carbohydrate chains was deduced to be six to eight sugar residues.
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002241:Carbohydrates / Q000032:analysis; D055598:Chemical Phenomena; D002621:Chemistry; D006023:Glycoproteins / Q000302:isolation & purification*; D007421:Intestine, Small / Q000032:analysis*; D009093:Mucus / Q000032:analysis*; D011485:Protein Binding; D013552:Swine; D014461:Ultracentrifugation; D014783:Viscosity
D000596:Amino Acids; D002241:Carbohydrates; D006023:Glycoproteins