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Detecting Hyaluronidase and Hyaluronidase Inhibitors

Hyaluronan-Substrate Gel and -Inverse Substrate Gel Techniques
  • Kazuhiro Mio
  • Antonei B. Csóka
  • Susan Nawy Stair
  • Robert Stern
Part of the Methods in Molecular Biology™ book series (MIMB, volume 171)

Abstract

Hyaluronidases are a group of enzymes that degrade the glycosaminoglycan hyaluronan (HA, hyaluronic acid). Many types of hyaluronidases are reported, from prokaryotes to eukaryotes (1,2). These enzymes have a wide variety of properties, including substrate specificity, inhibitor sensitivity, and a range of pH optima. Streptomyces hyaluronidase, and the venom hyaluronidases from bee, snake, and scorpion are active at neutral pH. Hyal-1, the best-characterized somatic hyaluronidase (3, 4, 5), product of one of the six hyaluronidase-like sequences in the human genome (6), is an acid-active enzyme with an optimum at pH 3.7. The sperm-specific PH-20 (7) has apparently two pH optima, pH 4.5 and 7.5, resulting possibly from two forms of the enzyme, membrane-bound and soluble (8,9)

Keywords

Sodium Dodecyl Sulfate Alcian Blue Coomassie Blue Staining Alcian Blue Staining Bromphenol Blue 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

References

  1. 1.
    Kreil, G. (1995) Hyaluronidases-a group of neglected enzymes. Protein Sci. 4,1666–1669.PubMedCrossRefGoogle Scholar
  2. 2.
    Csóka, T. B., Frost, G. I., and Stern, R. (1997) Hyaluronidases in tissue invasion. Invasion Metastasis 17, 297–311.PubMedGoogle Scholar
  3. 3.
    Frost, G. I., Csóka, T. B., Wong, T., and Stern, R. (1997) Purification, cloning, and expression of human plasma hyaluronidase. Biochem. Biophys. Res. Commun. 236, 10–15.PubMedCrossRefGoogle Scholar
  4. 4.
    Csóka T.B., Frost G.I., Wong T., and Stern R. (1997) Purification and microsequencing of hyaluronidase isozymes from human urine. FEBS Lett. 417, 307–310.PubMedCrossRefGoogle Scholar
  5. 5.
    Csóka, T. B., Frost, G. I., Heng, H. H. Q., Scherer, S. W., Mohapatra G., and Stern R. (1998) The hyaluronidase gene HYAL1 maps to chromosome 3p21.2-3p21.3 in human and 9F1-F2 in mouse, a conserved candidate tumor suppressor locus. Genomics 48, 63–70.PubMedCrossRefGoogle Scholar
  6. 6.
    Csóka, A. B., Scherer, S. W., and Stern, R. (1999) Expression analysis of six paralogous human hyaluronidase genes clustered on chromosomes 3p21 and 7q31. Genomics 60, 356–361.PubMedCrossRefGoogle Scholar
  7. 7.
    Gmachl, M., Sagan, S., Ketter, S., and Kreil, G. (1993) The human sperm protein PH-20 has hyaluronidase activity. FEBS Lett. 336, 545–548.PubMedCrossRefGoogle Scholar
  8. 8.
    Cherr, G. N., Meyers, S. A., Yudin, A. I., VandeVoort, C. A., Myles, D. G., Primakoff, P., and Overstreet, J. W. (1996) The PH-20 protein in Cynomolgus macaque spermatozoa: identification of two different forms exhibiting hyaluronidase activity. Develop. Biol. 175, 142–153.PubMedCrossRefGoogle Scholar
  9. 9.
    Meyer, M. F., Kreil, G., and Aschbauer, H. (1997) The soluble hyaluronidase from bull testes is a fragment of the membrane-bound PH-20 enzyme. FEBS Lett. 413, 385–388.PubMedCrossRefGoogle Scholar
  10. 10.
    Haas, E. (1946) On the mechanism of invasion. I. Antinvasin I, An enzyme in plasma, J. Biol. Chem. 163, 63–88.PubMedGoogle Scholar
  11. 11.
    Dorfman, A., Ott, M. L., and Whitney, R. (1948) The hyaluronidase inhibitor of human blood. J. Biol. Chem., 223, 621–629. 397.Google Scholar
  12. 12.
    Moore, D. H. and Harris, T. N. (1949) Occurrence of hyaluronidase inhibitors in fractions of electrophoretically separated serum. J. Biol. Chem. 179, 377–381.PubMedGoogle Scholar
  13. 13.
    Guntenhoener, M. W., Pogrel, M. A., and Stern, R. (1992) A substrate-gel assay for hyaluronidase activity. Matrix 12, 388–396.Google Scholar
  14. 14.
    Miura, R. O., Yamagata, S., Miura, Y., Harada, T., and Yamagata, T. (1995) Analysis of glycosaminoglycan-degrading enzymes by substrate gel electrophoresis (zymography). Anal Biochem. 225, 333–340.PubMedCrossRefGoogle Scholar
  15. 15.
    Mio, K., Carette, O., Maibach, H. I., and Stern, R. (2000) A serum inhibitor of hyaluronidase. J. Biol. Chem., July 24.Google Scholar
  16. 16.
    Laemmli, UK. (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227, 680–685.Google Scholar
  17. 17.
    Wardi, A. H. and Michos, G. A. (1972) Alcian blue staining of glycoproteins in acrylamide disc electrophoresis. Anal. Biochem. 49, 607–609.PubMedCrossRefGoogle Scholar
  18. 18.
    Cowman, M. K., Slahetka, M. F., Hittner, D. M., Kim, J., Forino, M., and Gadelrab, G. (1984) Polyacrylamide-gel electrophoresis and Alcian blue staining of sulphated glycosaminoglycan oligosaccharides. Biochem. J. 221, 707–716.PubMedGoogle Scholar
  19. 19.
    Turner, R. E. and Cowman, M. K. (1985) Cationic dye binding by hyaluronate fragments: dependence on hyaluronate chain length. Arch. Biochem. Biophys. 237, 253–260.PubMedCrossRefGoogle Scholar
  20. 20.
    Wall, R. S. and Gyi, T. J. (1988) Alcian blue staining of proteoglycans in polyacrylamide gels using the &quote;critical electrolyte concentration&quote; approach. Anal. Biochem. 175, 298–299.PubMedCrossRefGoogle Scholar
  21. 21.
    Ghiggeri, G. M., Candiano, G., Ginevri, F., Mutti, A., Bergamaschi, E., Alinovi, R., and Righetti, P. G. (1988) Hydrophobic interaction of Alcian blue with soluble and erythrocyte membrane proteins. J. Chromatogr. 452, 347–357.PubMedCrossRefGoogle Scholar
  22. 22.
    Johnston, J. P. (1955) The sedimentation behavior of mixtures of hyaluronic acid and albumin in the ultracentrifuge. Biochem. J. 59, 620–627.PubMedGoogle Scholar
  23. 23.
    Davies, M., Nichol, L. W., and Ogston, A. G. (1963) Frictional effects in the migration of mixtures of hyaluronic acid and serum albumin. Biochim. Biophys. Acta 75, 436–438.PubMedCrossRefGoogle Scholar
  24. 24.
    Gramling, E., Niedermeier, W., Holley, H. L., and Pigman, W. (1963) Some factors affecting the interaction of hyaluronic acid with bovine plasma albumin. Biochim. Biophys.Acta 69, 552–558.PubMedCrossRefGoogle Scholar

Copyright information

© Humana Press Inc., Totowa, NJ 2001

Authors and Affiliations

  • Kazuhiro Mio
  • Antonei B. Csóka
  • Susan Nawy Stair
  • Robert Stern

There are no affiliations available

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