Hostname: page-component-848d4c4894-hfldf Total loading time: 0 Render date: 2024-05-29T15:50:45.463Z Has data issue: false hasContentIssue false
  • English
  • Français

Purification and characterization of a novel 3′-5′ DNA helicase from Plasmodium falciparum and its sensitivity to anthracycline antibiotics

Published online by Cambridge University Press:  13 June 2006

P. SUNTORNTHITICHAROEN ,
S. PETMITR  and
P. CHAVALITSHEWINKOON-PETMITR
P. SUNTORNTHITICHAROEN
Affiliation:
Department of Protozoology, Faculty of Tropical Medicine, Mahidol University, 420/6 Ratchawithi Road, Bangkok 10400, Thailand
S. PETMITR
Affiliation:
Department of Tropical Nutrition and Food Science, Faculty of Tropical Medicine, Mahidol University, 420/6 Ratchawithi Road, Bangkok 10400, Thailand
P. CHAVALITSHEWINKOON-PETMITR
Affiliation:
Department of Protozoology, Faculty of Tropical Medicine, Mahidol University, 420/6 Ratchawithi Road, Bangkok 10400, Thailand
Get access Rights & Permissions [Opens in a new window]

Abstract

Plasmodium falciparum has developed resistance to most anti-malarials; therefore, an investigation of potential targets should be performed. DNA helicases are enzymes that catalyse the unwinding of double-stranded DNA to provide single-stranded templates for DNA replication, repair and recombination. In this study, a DNA helicase (PfDH A) was purified from a crude extract of Plasmodium falciparum. DNA helicase activity was measured by assaying unwinding activity. The apparent molecular weight of PfDH A as determined by SDS-PAGE was 90 kDa. PfDH A moved unidirectionally in the 3′ -to- 5′ direction along the bound strand and preferred a fork-like substrate structure and could not unwind blunt-ended duplex DNA. Unwinding activity required Mg2+ and could be inhibited by 200 mM NaCl or KCl and was dependent on hydrolysis of ATP or dATP. Anthracyclines, including daunorubicin, nogalamycin, doxorubicin, and aclarubicin, inhibited PfDH A activity with IC50 values of 2, 5, 8 and 9 μM, respectively. Based on the results, PfDH A differs from all known human DNA helicases. However, its function and roles in parasite DNA replication need to be elucidated in the future.

Keywords

malariaPlasmodium falciparumDNA helicaseDNA-interacting agents
Type
Research Article
Information
Parasitology , Volume 133 , Issue 4 , October 2006 , pp. 389 - 398
Copyright
© 2006 Cambridge University Press

Access options

Get access to the full version of this content by using one of the access options below. (Log in options will check for institutional or personal access. Content may require purchase if you do not have access.)

References

REFERENCES

Bachur, N. R., Yu, F., Johnson, R., Hickey, R., Wu, Y. and Malkas, L. ( 1992). Helicase inhibition by anthracycline anticancer agents. Molecular Pharmacology 41, 993–998.Google Scholar
Bachur, N. R., Johnson, R., Yu, F., Hickey, R., Applegren, N. and Malkas, L. ( 1993). Antihelicase action of DNA-binding anticancer agents: relationship to guanosine-cytidine intercalator binding. Molecular Pharmacology 44, 1064–1069.Google Scholar
Chandra, P., Zunino, F., Gotz, A., Gericke, D., Thorbeck, R and Di Marco, A. ( 1972). Specific inhibition of DNA-polymerases from RNA tumor viruses by some new daunomycin derivatives. FEBS Letters 21, 264–268.CrossRefGoogle Scholar
Chavalitshewinkoon, P. and Wilairat, P. ( 1991). A simple technique for large scale in vitro culture of Plasmodium falciparum. Southeast Asian Journal of Tropical Medicine and Public Health 22, 544–547.Google Scholar
Ciarrocchi, G., Lestingi, M., Fontana, M., Spadari, S. and Montecucco, A. ( 1991). Correlation between anthracycline structure and human DNA ligase inhibition. The Biochemical Journal 279, 141–146.CrossRefGoogle Scholar
Costa, M., Ochem, A., Staub, A. and Falaschi, A. ( 1999). Human DNA helicase VIII: a DNA and RNA helicase corresponding to the G3BP protein, an element of the ras transduction pathway. Nucleic Acids Research 27, 817–821.CrossRefGoogle Scholar
Goetz, G. S., Dean, F., Hurwitz, J. and Matson, S. W. ( 1988). The unwinding of duplex regions in DNA by the simian virus 40 large tumor antigen-associated DNA helicase activity. Journal of Biological Chemistry 263, 383–392.Google Scholar
Goodman, M. F., Bessman, M. J. and Bachur, N. R. ( 1974). Adriamycin and daunorubicin inhibition of mutant T4 DNA polymerases. Proceedings of the National Academy of Sciences, USA 71, 1193–1196.CrossRefGoogle Scholar
Hartmann, G., Goller, H., Koschel, K, Kersten, W. and Kersten, H. ( 1964). Inhibition of DNA-dependent RNA and DNA synthesis by antibiotics. Biochemische Zeitschrift 341, 126–128.Google Scholar
Lambros, C. and Vanderberg, J. P. ( 1979). Synchronization of Plasmodium falciparum erythrocytic stages in culture. Journal of Parasitology 65, 418–420.CrossRefGoogle Scholar
Lee, Y. C., Byfield, J., Bennett, L. R. and Chan, Y. M. ( 1974). X-ray repair replication in L1210 leukemia cells. Cancer Research 34, 2624–2633.Google Scholar
Lohman, T. M. and Bjornson, K. P. ( 1996). Mechanisms of helicase-catalyzed DNA unwinding. Annual Review of Biochemistry 65, 169–214.CrossRefGoogle Scholar
Matson, S. W. and Kaiser-Rogers, K. A. ( 1990). DNA helicases. Annual Review of Biochemistry 59, 289–329.CrossRefGoogle Scholar
Matson, S. W. and Bean, D. W. ( 1995). Purification and biochemical characterization of enzymes with DNA helicase activity. Methods in Enzymology 262, 389–405.CrossRefGoogle Scholar
Pradhan, A., Chauhan, V. S. and Tuteja, R. ( 2005 a). Plasmodium falciparum DNA helicase 60 is a schizont stage specific, bipolar and dual helicase stimulated by PKC phosphorelation. Molecular and Biochemical Parasitology 144, 133–141.Google Scholar
Pradhan, A., Chauhan, V. S. and Tuteja, R. ( 2005 b). A novel ‘DEAD-box’ DNA helicase from Plasmodium falciparum is homologous to p68. Molecular and Biochemical Parasitology 140, 55–60.Google Scholar
Runyon, G. T. and Lohman, T. M. ( 1989). Escherichia coli helicase II (uvrD) protein can completely unwind fully duplex linear and nicked circular DNA. Journal of Biological Chemistry 264, 17502–17512.Google Scholar
Sachs, J. and Malaney, P. ( 2002). The economic and social burden of malaria. Nature, London 415, 680–685.CrossRefGoogle Scholar
Seki, M. and Enomoto, T., Hanaoka, F. and Yamada, M. ( 1987). DNA-dependent adenosinetriphosphatase B from mouse FM3A cells has DNA helicase activity. Biochemistry 26, 2924–2928.CrossRefGoogle Scholar
Stahl, H. and Knippers, R. ( 1987). The simian virus 40 large tumor antigen. Biochimica et Biophysica Acta 910, 1–10.CrossRefGoogle Scholar
Tewey, K. M., Rowe, T., Yang, L., Halligan, B. D. and Liu, L. F. ( 1984). Adriamycin-induced DNA damage mediated by mammalian DNA topoisomerase II. Science 226, 466–468.CrossRefGoogle Scholar
Thaithong, S., Beale, G. and Chutmongkonkul, M. ( 1983). Susceptibility of Plasmodium falciparum to five drugs: an in vitro study of isolates mainly from Thailand. Transactions of the Royal Society of Tropical Medicine and Hygiene 77, 228–231.CrossRefGoogle Scholar
Thommes, P. and Hubscher, U. ( 1992). Eukaryotic DNA helicases: essential enzymes for DNA transactions. Chromosoma 101, 467–473.CrossRefGoogle Scholar
Tuteja, N., Tuteja, R., Rahman, K., Kang, L. Y. and Falaschi, A. ( 1990). A DNA helicase from human cells. Nucleic Acids Research 18, 6785–6792.CrossRefGoogle Scholar
Tuteja, N., Rahman, K., Tuteja, R., Ochem, A., Skopac, D. and Falaschi, A. ( 1992). DNA helicase III from HeLa cells: an enzyme that acts preferentially on partially unwound DNA duplexes. Nucleic Acids Research 20, 5329–5337.CrossRefGoogle Scholar
Tuteja, N., Rahman, K., Tuteja, R. and Falaschi, A. ( 1993). Human DNA helicase V, a novel DNA unwinding enzyme from HeLa cells. Nucleic Acids Research 21, 2323–2329.CrossRefGoogle Scholar
Tuteja, N., Tuteja, R., Ochem, A., Taneja, P., Huang, N. W., Simaocsits, A., Susic, S., Rahman, K., Marusic, L., Chen, J., Zhang, J., Wang, S., Pongor, S. and Falaschi, A. ( 1994). Human DNA helicase II: a novel DNA unwinding enzyme identified as the Ku autoantigen. EMBO Journal 13, 4991–5001.Google Scholar
Tuteja, N., Ochem, A., Taneja, P., Tuteja, R., Skopac, D. and Falaschi, A. ( 1995). Purification and properties of human DNA helicase VI. Nucleic Acids Research 23, 2457–2463.CrossRefGoogle Scholar
Tuteja, R., Malhotra, P., Song, P., Tuteja, N. and Chauhan, V. S. ( 2002). Isolation and characterization of an eIF-4A homologue from Plasmodium cynomolgi. Molecular and Biochemical Parasitology 124, 79–83.CrossRefGoogle Scholar
Tuteja, R., Tuteja, N., Malhotra, P. and Chauhan, V. S. ( 2003). Replication fork-stimulated eIF-4A from Plasmodium cynomolgi unwinds DNA in the 3′ to 5′ direction and is inhibited by DNA-interacting compounds. Archives of Biochemistry and Biophysics 414, 108–114.CrossRefGoogle Scholar
Vindigni, A., Ochem, A., Triolo, G. and Falaschi, A. ( 2001). Identification of human DNA helicase V with the far upstream element-binding protein. Nucleic Acids Research 29, 1061–1067.CrossRefGoogle Scholar
Zhang, S. S. and Grosse, F. ( 1991). Purification and characterization of two DNA helicases from calf thymus nuclei. Journal of Biological Chemistry 266, 20483–20490.Google Scholar