Skip to main content
SpringerLink
Log in

Conformational Stability and Hemolytic Activity of Actinoporin RTX-SII from the Sea Anemone Radianthus macrodactylus

  • Published:
Biochemistry (Moscow) Aims and scope Submit manuscript

Abstract

The spatial organization of actinoporin RTX-SII from the sea anemone Radianthus macrodactylus on the level of tertiary and secondary structures was studied by UV and CD spectroscopy and intrinsic protein fluorescence. The specific and molar extinction coefficients of RTX-SII were determined. The percentages of canonical secondary structures of actinoporin were calculated. The tertiary structure of the polypeptide is well developed and its secondary structure is highly ordered and contains about 50% antiparallel folded β-sheets. The irreversible thermal denaturation of RTX-SII was studied by CD spectroscopy; a conformational transition occurs at 53°C. Above this temperature irreversible conformational changes are observed in the secondary and tertiary structures. This is accompanied by redistribution of the content of regular and distorted forms of β-sheet and also by increase in the content of an unordered form. It is suggested that an intermediate is formed in the process of thermal denaturation. Acid-base titration of RTX-SII results in irreversible conformational changes at pH below 2.0 and above 12.0. As shown by intrinsic protein fluorescence, tyrosine residues of RTX-SII make a fundamental contribution to emission, and the total fluorescence depends more on temperature and ionic strength of the solution than tryptophan fluorescence. The data on conformational stability of actinoporin are correlated with data on its hemolytic activity. Activity of RTX-SII significantly decreases at increased temperature and slightly decreases at low pH. Hemolytic activity drastically increases at high pH. Increase in the actinoporin activity at pH above 10 seems to be caused by ionization of the molecule.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

REFERENCES

  1. Norton, R. S. (1998) J. Toxicol.-Toxin Rev., 17, 99–130.

    Google Scholar 

  2. Macek, P. (1992) Microbiol. Immunol., 105, 121–130.

    Article  Google Scholar 

  3. Turk, T. (1991) J. Toxicol.-Toxin Rev., 10, 223–262.

    Google Scholar 

  4. Galettis, P., and Norton, R. S. (1990) Toxicon, 28, 695–706.

    Article  PubMed  Google Scholar 

  5. Batista, U., Macek, P., and Sedmak, B. (1986) Period. Biol., 88, 97–98.

    Google Scholar 

  6. Tejica, M., Anderluh, G., Macek, P., Marcet, R., Torres, D., Sarracent, J., Alvarez, C., Lanio, M. E., Serra, M. D., and Menestrina, G. (1999) Int. J. Parasitol., 29, 489–498.

    Article  PubMed  Google Scholar 

  7. Chanturiya, A. N. (1990) Biochim. Biophys. Acta, 1026, 248–250.

    PubMed  Google Scholar 

  8. Shnyrov, V. L., Monastyrnaya, M. M., Zhadan, G. G., Kuznetsova, S. M., and Kozlovskaya, E. P. (1992) Biochem. Int., 26, 219–229.

    PubMed  Google Scholar 

  9. Zhadan, G. G., Kuznetsova, S. M., Opalikova, O. V., Monastyrnaya, M. M., Zykova, T. A., Emelyanenko, V. I., Villar, E., and Shnyrov, V. L. (1994) Biochem. Mol. Biol. Int., 32, 331–340.

    PubMed  Google Scholar 

  10. Anderluh, G., and Macek, P. (2002) Toxicon, 40, 111–124.

    Article  PubMed  Google Scholar 

  11. Anderluh, G., Krizaj, I., Strukelj, B., Gubensek, F., Macek, P., and Pungercar, J. (1999) Toxicon, 37, 1391–1401.

    Article  PubMed  Google Scholar 

  12. Norton, R. S., Bobek, G., Ivanov, J. O., Thomson, M., Fiala-Beer, E., Moritz, R. L., and Simpson, R. J. (1990) Toxicon, 28, 29–41.

    Article  PubMed  Google Scholar 

  13. Huerta, V., Morera, V., Guanche, Y., Cinea, G., Gonzales, L. J., Betancourt, L., Martinez, D., Alvarez, C., Lanio, M. E., and Besada, V. (2001) Toxicon, 39, 1253–1256.

    Article  PubMed  Google Scholar 

  14. Khoo, K. S., Kam, W. K., Khoo, H. E., Gopalakrishnakone, P., and Chung, M. C. M. (1993) Toxicon, 31, 1567–1579.

    Article  PubMed  Google Scholar 

  15. Monastyrnaya, M. M., Zykova, T. A., Apalikova, O. W., Shwets, T. W., and Kozlovskaya, E. P. (2002) Toxicon, 40, 1197–1217.

    Article  PubMed  Google Scholar 

  16. Klyshko, E. V., Il’yna, A. P., Likhatskaya, G. N., Issaeva, M. P., Guzev, K. V., Monastyrnaya, M. M., Kozlovskaya, E. P., Lipkin, A. V., Barsova, E. I., Kryzko, E. V., Trifonov, E. V., and Nurminsky, E. A. (2004) Vestnik DVO RAN, 3, 45–53.

    Google Scholar 

  17. Klyshko, E. V., Issaeva, M. P., Monastyrnaya, M. M., Il’yna, A. P., Guzev, K. V., Vakorina, T. I., Dmitrenok, P. S., Zykova, T. A., and Kozlovskaya, E. P. (2004) Toxicon, 44, 315–326.

    Article  PubMed  Google Scholar 

  18. Lowry, O. H., Rosebrough, N. J., Farr, A. L., and Randall, R. J. (1951) J. Biol. Chem., 193, 265–275.

    PubMed  Google Scholar 

  19. Kitagava, I., Nishino, T., Kobayashi, M., and Kyogoku, Y. (1981) Chem. Pharm. Bull., 29, 1951–1956.

    Google Scholar 

  20. Winder, A. F., and Gent, W. L. G. (1971) Biopolymers, 10, 1243–1251.

    Article  PubMed  Google Scholar 

  21. Ichikawa, T., and Johnson, C. Yr. (1983) Chem. Pharm. Bull., 29, 438–444.

    Google Scholar 

  22. Provencher, S. W., and Glockner, W. C. (1981) Biochemistry, 20, 33–37.

    Article  PubMed  Google Scholar 

  23. Belmonte, G., Menestrina, G., Pederzolli, C., Krizaj, I., Gubensek, F., Turk, T., and Macek, P. (1994) Biochim. Biophys. Acta, 1192, 197–204.

    PubMed  Google Scholar 

  24. Poklar, N., Lah, J., Salobir, M., Macek, P., and Vesnaver, G. (1997) Biochemistry, 36, 14345–14352.

    Article  PubMed  Google Scholar 

  25. Menestrina, G., Cabiaux, V., and Tejuca, M. (1999) Biochem. Biophys. Res. Commun., 254, 174–180.

    Article  PubMed  Google Scholar 

  26. Athanasiadis, A., Anderluh, G., Macek, P., and Turk, D. (2001) Structure, 9, 341–346.

    Article  PubMed  Google Scholar 

  27. Mancheno, J. M., Martin-Benito, J., Martinez-Ripoll, M., Gavilanes, J. G., and Hermoso, J. A. (2003) Structure, 11, 1–20.

    PubMed  Google Scholar 

  28. Ven’yaminov, S. Yu., Kosykh, V. G., Kholodkov, O. A., and Bur’yanov, Ya. I. (1990) Bioorg. Khim., 16, 47–51.

    PubMed  Google Scholar 

  29. France, L. L., Kieleczawa, J. J., Dunn, J. J., Hind, G., and Sutherland, J. C. (1992) Biochim. Biophys. Acta, 1120, 59–68.

    PubMed  Google Scholar 

  30. Moroz, S. V., Glazunov, V. P., Vakorina, T. I., Pavlenko, A. F., Odinokov, S. E., and Ovodov, Yu. S. (1987) Bioorg. Khim., 13, 519–527.

    PubMed  Google Scholar 

  31. http:/lamar.colostate.edu/∼sreeram/CDPro

  32. Alvarez, C., Lanio M. E., Tejuca, M., Martinez, D., Pazos, F., Campos, A. M., Encinas, M. V., Pertinhez, T., Schreier, S., and Lissi, E. A. (1998) Toxicon, 36, 165–178.

    Article  PubMed  Google Scholar 

  33. Lakovich, D. (1986) Basics of Fluorescence Spectroscopy [Russian translation], Mir, Moscow.

    Google Scholar 

  34. Macek, P., Zecchini, M., Pederzolli, C., Serra, M. D., and Menestrina, G. (1995) Eur. J. Biochem., 234, 329–335.

    Article  PubMed  Google Scholar 

  35. Alvarez, C., Pazos, I. F., Lanio, M. E., Martinez, D., Schreier, S., Casallanovo, F., Campos, A. M., and Lissi, E. (2001) Toxicon, 39, 539–553.

    Article  PubMed  Google Scholar 

  36. Eftink, M. R. (1998) Biochemistry (Moscow), 63, 276–284.

    PubMed  Google Scholar 

  37. Doyle, J. W., and Kem, W. R. (1989) Biochim. Biophys. Acta, 987, 181–186.

    PubMed  Google Scholar 

  38. Meinardi, E., Azcurra, J. M., Florin-Christensen, M., and Florin-Christensen, J. (1994) Comp. Biochem. Physiol., 109B, 153–161.

    Google Scholar 

  39. Macek, P., and Lebez, D. (1981) Toxicon, 19, 233–240.

    Article  PubMed  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Pacific Institute of Bioorganic Chemistry, Far East Branch of the Russian Academy of Sciences, pr. 100-let Vladivostoku 159, 690022, Vladivostok, Russia

    T. I. Vakorina, E. V. Klyshko, M. M. Monastyrnaya & E. P. Kozlovskaya

Authors
  1. T. I. Vakorina
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. E. V. Klyshko
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. M. M. Monastyrnaya
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. E. P. Kozlovskaya
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to T. I. Vakorina.

Additional information

__________

Translated from Biokhimiya, Vol. 70, No. 7, 2005, pp. 957–967.

Original Russian Text Copyright © 2005 by Vakorina,, Klyshko, Monastyrnaya, Kozlovskaya.

Originally published in Biochemistry (Moscow) On-Line Papers in Press, as Manuscript BM04-164, December 5, 2004.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Vakorina, T.I., Klyshko, E.V., Monastyrnaya, M.M. et al. Conformational Stability and Hemolytic Activity of Actinoporin RTX-SII from the Sea Anemone Radianthus macrodactylus . Biochemistry (Moscow) 70, 790–798 (2005). https://doi.org/10.1007/s10541-005-0185-1

Download citation

  • Received:

  • Revised:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10541-005-0185-1

Key words

Search

Navigation