Abstract
Destabilase, endo-ε-(γ-Glu)-Lys-isopeptidase, was prepared from the salivary gland secretion of the medicinal leech (Hirudo medicinalis). The secretion prepared by the known method of Rigbi et al. (1987) (secretion-K) lacks the destabilase-characteristic highly specific isopeptidase activity (the D-dimer-monomerizing activity) because of its degradation by proteolytic activity (the substrate of Glp-Ala-Ala-Leu-pNA) due to contamination with leech intestinal channel contents. Therefore, we have elaborated a new technique for preparation of a true leech secretion (secretion-I). This secretion is characterized by the complete absence of the leech intestinal channel contents and has no proteolytic activity. For the first time the destabilase-specific D-dimer-monomerizing and lysozyme activities were separated by fractionation of secretion-I by HPLC gel filtration through Superose S-12. For the purified destabilase preparation, these activities were separated by reversed-phase chromatography in an acetonitrile gradient (0-60%) in the presence of 0.1% trifluoroacetic acid. The monomerizing activity of destabilase is responsible for the ability of secretion-I to dissolve stabilized fibrin via isopeptidolysis of α-α and γ-γ fibrin chains bound by ε-(γ-Glu)-Lys-isopeptide bonds.
Similar content being viewed by others
REFERENCES
Baskova, I. P., and Nikonov, G. I. (1985) Biokhimiya, 50, 424–431.
Zavalova, L. L., Kuzina, E. V., Levina, N. B., and Baskova, I. P. (1993) Thromb. Res., 71, 241–244.
Zavalova, L., Lukyanov, S., Baskova, I., Snezhkov, E., Berezhnoy, S., Bogdanova, E., Barsova, E., and Sverdlov, E. (1996) Mol. Gen. Genet., 253, 20–25.
Aeschlimann, D., and Paulsson, M. (1994) Thromb. Haemostasis, 71, 402–415.
Zavalova, L. L., Baskova, I. P., Lukyanov, S. A., Sass, A. V., Snezhkov, E. V., Akopov, S. B., Artamonova, I. I., Archipova, V. S., Nesmeyanov, V. A., Kozlov, D. G., Benevolensky, S. V., Kiseleva, V. I., Poverenny, A. M., and Sverdlov, E. D. (2000) Biochim. Biophys. Acta, 1487, 69–77.
Rigbi, M., Levy, H., Iraquy, F., Teitelbaum, M., Orevi, M., Alajoutsijarvi, A., Horovitz, A., and Galun, R. (1987) Comp. Biochem. Physiol. [B], 87, 563–573.
Belitser, V. A., Pozdnjakova, T. M., Platonova, T. N., and Vovk, E. V. (1981) Thromb. Res., 21, 565–572.
Lyublinskaya, L. A., Khaidu, I., Balandina, G. N., Filippova, I. Yu., Markaryan, A. N., Lysogorskaya, E. N., Oksenoit, E. S., and Stepanov, V. M. (1987) Bioorg. Khim., 13, 748–753.
Harlow, E., and Lane, D. (1988) Antibodies. A Laboratory Manual, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York, pp. 479–511.
Jespersen, J., and Astrup, T. (1983) Haemostasis, 13, 301–315.
Sedmak, J. J., and Grossberg, S. E. (1977) Analyt. Biochem., 79, 544–552.
Laemmly, U. K. (1970) Nature, 227, 680–685.
Budzynski, A. Z., Marder, V. J., and Shainoff, J. R. (1974) J. Biol. Chem., 249, 2294–2302.
Baskova, I. P. (1994) The Method for Preparation of Bloodsucking Leech Salivary Gland Secretion, Patent No. 2045954.
Romanova, T. V., Rudenskaya, G. N., Tebloeva, L. T., Zverev, D. V., and Emirova, Kh. M. (1998) Vopr. Med. Khim., 44, 296–304.
Baskova, I. P., and Nikonov, G. I. (1984) Folia Haematol., 111, 131–137.
Baskova, I. P., Khalil, S., Nikonov, G. I., Rabinovich, S. E., Nartikova, N. F., and Paskhina, T. S. (1989) Biokhimiya, 53, 1467–1473.
Baskova, I. P., Nikonov, G. I., and Khalil, S. (1988) Folia Haematol., 115, 166–170.
Dalbey, R. I., and von Heijne, G. (1992) Trends Biochem. Sci., 17, 474–478.
Baskova, I. P., Nikonov, G. I., Zavalova, L. L., and Larionova, N. I. (1990) Biokhimiya, 55, 674–679.
Baskova, I. P., Zavalova, L. L., and Kuzina, E. V. (1994) Bioorg. Khim., 20, 492–497.
Medved, L. V., Gorkun, O. V., and Privalov, P. L. (1983) FEBS Lett., 160, 291–295.
Zavalova, L. L., Nikonov, G. I., Popova, G. Yu., and Baskova, I. P. (1991) Biokhimiya, 56, 115–124.
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Baskova, I.P., Zavalova, L.L., Basanova, A.V. et al. Separation of Monomerizing and Lysozyme Activities of Destabilase from Medicinal Leech Salivary Gland Secretion. Biochemistry (Moscow) 66, 1368–1373 (2001). https://doi.org/10.1023/A:1013333829196
Issue Date:
DOI: https://doi.org/10.1023/A:1013333829196