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. 1981 Apr 1;195(1):267–275. doi: 10.1042/bj1950267

Isolation and characterization of the native glycoprotein from pig small-intestinal mucus.

M Mantle, A Allen
PMCID: PMC1162882  PMID: 7306053

Abstract

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.

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Selected References

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  1. AMINOFF D. Methods for the quantitative estimation of N-acetylneuraminic acid and their application to hydrolysates of sialomucoids. Biochem J. 1961 Nov;81:384–392. doi: 10.1042/bj0810384. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Allen A., Pain R. H., Robson T. R. Model for the structure of the gastric mucous gel. Nature. 1976 Nov 4;264(5581):88–89. doi: 10.1038/264088a0. [DOI] [PubMed] [Google Scholar]
  3. Allen A. Structure of gastrointestinal mucus glycoproteins and the viscous and gel-forming properties of mucus. Br Med Bull. 1978 Jan;34(1):28–33. [PubMed] [Google Scholar]
  4. Bara J., Paul-Gardais A., Loisillier F., Burtin P. Isolation of a sulfated glycopeptidic antigen from human gastric tumors: its localization in normal and cancerous gastrointestinal tissues. Int J Cancer. 1978 Feb 15;21(2):133–139. doi: 10.1002/ijc.2910210202. [DOI] [PubMed] [Google Scholar]
  5. Bella A., Jr, Kim Y. S. Rat small intestinal mucin: isolation and characterization of a water-soluble mucin fraction. Arch Biochem Biophys. 1972 Jun;150(2):679–689. doi: 10.1016/0003-9861(72)90086-0. [DOI] [PubMed] [Google Scholar]
  6. Carlson D. M. Structures and immunochemical properties of oligosaccharides isolated from pig submaxillary mucins. J Biol Chem. 1968 Feb 10;243(3):616–626. [PubMed] [Google Scholar]
  7. Chambers R. E., Clamp J. R. An assessment of methanolysis and other factors used in the analysis of carbohydrate-containing materials. Biochem J. 1971 Dec;125(4):1009–1018. doi: 10.1042/bj1251009. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Clarke A. E., Denborough M. A. The interaction of concanavalin A with blood-group-substance glycoproteins from human secretions. Biochem J. 1971 Mar;121(5):811–816. doi: 10.1042/bj1210811. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Creeth J. M. Constituents of mucus and their separation. Br Med Bull. 1978 Jan;34(1):17–24. doi: 10.1093/oxfordjournals.bmb.a071454. [DOI] [PubMed] [Google Scholar]
  10. Creeth J. M., Denborough M. A. The use of equilibrium-density-gradient methods for the preparation and characterization of blood-group-specific glycoproteins. Biochem J. 1970 May;117(5):879–891. doi: 10.1042/bj1170879. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Creeth J. M., Pain R. H. The determination of molecular weights of biological macromolecules by ultracentrifuge methods. Prog Biophys Mol Biol. 1967;17:217–287. doi: 10.1016/0079-6107(67)90008-9. [DOI] [PubMed] [Google Scholar]
  12. Forstner J. F., Jabbal I., Forstner G. G. Goblet cell mucin of rat small intestine. Chemical and physical characterization. Can J Biochem. 1973 Aug;51(8):1154–1166. doi: 10.1139/o73-152. [DOI] [PubMed] [Google Scholar]
  13. Forstner J. F., Jabbal I., Qureshi R., Kells D. I., Forstner G. G. The role of disulphide bonds in human intestinal mucin. Biochem J. 1979 Sep 1;181(3):725–732. doi: 10.1042/bj1810725. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Jabbal I., Kells D. I., Forstner G., Forstner J. Human intestinal goblet cell mucin. Can J Biochem. 1976 Aug;54(8):707–716. doi: 10.1139/o76-102. [DOI] [PubMed] [Google Scholar]
  15. List S. J., Findlay B. P., Forstner G. G., Forstner J. F. Enhancement of the viscosity of mucin by serum albumin. Biochem J. 1978 Nov 1;175(2):565–571. doi: 10.1042/bj1750565. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Mantle M., Allen A. A colorimetric assay for glycoproteins based on the periodic acid/Schiff stain [proceedings]. Biochem Soc Trans. 1978;6(3):607–609. doi: 10.1042/bst0060607. [DOI] [PubMed] [Google Scholar]
  17. Mantle M., Allen A. Isolation of the major water-soluble glycoprotein from pig small-intestine mucus and evidence for relatively simple carbohydrate side chains [proceedings]. Biochem Soc Trans. 1979 Apr;7(2):393–395. doi: 10.1042/bst0070393. [DOI] [PubMed] [Google Scholar]
  18. Mantle M., Mantle D., Allen A. Polymeric structure of pig small-intestinal mucus glycoprotein. Dissociation by proteolysis or by reduction of disulphide bridges. Biochem J. 1981 Apr 1;195(1):277–285. doi: 10.1042/bj1950277. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Marshall T., Allen A. The isolation and characterization of the high-molecular-weight glycoprotein from pig colonic mucus. Biochem J. 1978 Aug 1;173(2):569–578. doi: 10.1042/bj1730569. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Pearson J., Allen A., Venables C. Gastric mucus: isolation and polymeric structure of the undegraded glycoprotein: its breakdown by pepsin. Gastroenterology. 1980 Apr;78(4):709–715. [PubMed] [Google Scholar]
  21. Roberts G. P. The role of disulfide bonds in maintaining the gel structure of bronchial mucus. Arch Biochem Biophys. 1976 Apr;173(2):528–537. doi: 10.1016/0003-9861(76)90289-7. [DOI] [PubMed] [Google Scholar]
  22. Sajdera S. W., Hascall V. C. Proteinpolysaccharide complex from bovine nasal cartilage. A comparison of low and high shear extraction procedures. J Biol Chem. 1969 Jan 10;244(1):77–87. [PubMed] [Google Scholar]
  23. Scawen M., Allen A. The action of proteolytic enzymes on the glycoprotein from pig gastric mucus. Biochem J. 1977 May 1;163(2):363–368. doi: 10.1042/bj1630363. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Slomiany B. L., Meyer K. Isolation and structural studies of sulfated glycoproteins of hog gastric mucosa. J Biol Chem. 1972 Aug 25;247(16):5062–5070. [PubMed] [Google Scholar]
  25. Snary D., Allen A., Pain R. H. Conformational changes in gastric mucoproteins induced by caesium chloride and guanidinium chloride. Biochem J. 1974 Sep;141(3):641–646. doi: 10.1042/bj1410641. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Snary D., Allen A., Pain R. H. The structure of pig gastric mucus. Conformational transitions induced by salt. Eur J Biochem. 1971 Dec 22;24(1):183–189. doi: 10.1111/j.1432-1033.1971.tb19669.x. [DOI] [PubMed] [Google Scholar]
  27. Snary D., Allen A. Studies on gastric mucoproteins. The isolation and characterization of the mucoprotein of the water-soluble mucus from pig cardiac gastric mucosa. Biochem J. 1971 Aug;123(5):845–853. doi: 10.1042/bj1230845. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Starkey B. J., Snary D., Allen A. Characterization of gastric mucoproteins isolated by equilibrium density-gradient centrifugation in caesium chloride. Biochem J. 1974 Sep;141(3):633–639. doi: 10.1042/bj1410633. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Watkins W. M. Blood-group substances. Science. 1966 Apr 8;152(3719):172–181. doi: 10.1126/science.152.3719.172. [DOI] [PubMed] [Google Scholar]
  30. Weber K., Pringle J. R., Osborn M. Measurement of molecular weights by electrophoresis on SDS-acrylamide gel. Methods Enzymol. 1972;26:3–27. doi: 10.1016/s0076-6879(72)26003-7. [DOI] [PubMed] [Google Scholar]
  31. YPHANTIS D. A. EQUILIBRIUM ULTRACENTRIFUGATION OF DILUTE SOLUTIONS. Biochemistry. 1964 Mar;3:297–317. doi: 10.1021/bi00891a003. [DOI] [PubMed] [Google Scholar]

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