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Molecular characterization of a putative protein disulfide isomerase from Babesia caballi

Published online by Cambridge University Press:  15 August 2005

H. IKADAI ,
R. TSUKADA ,
M. SASAKI ,
R. TAKASHIRO ,
N. YOKOYAMA ,
N. KUDO ,
I. IGARASHI  and
T. OYAMADA
H. IKADAI
Affiliation:
Department of Veterinary Parasitology, School of Veterinary Medicine and Animal Sciences, Kitasato University, Towada, Aomori 034-8628, Japan
R. TSUKADA
Affiliation:
Department of Veterinary Parasitology, School of Veterinary Medicine and Animal Sciences, Kitasato University, Towada, Aomori 034-8628, Japan
M. SASAKI
Affiliation:
Department of Veterinary Parasitology, School of Veterinary Medicine and Animal Sciences, Kitasato University, Towada, Aomori 034-8628, Japan
R. TAKASHIRO
Affiliation:
Department of Veterinary Parasitology, School of Veterinary Medicine and Animal Sciences, Kitasato University, Towada, Aomori 034-8628, Japan
N. YOKOYAMA
Affiliation:
National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Obihiro, Hokkaido 080-8555, Japan
N. KUDO
Affiliation:
Department of Veterinary Parasitology, School of Veterinary Medicine and Animal Sciences, Kitasato University, Towada, Aomori 034-8628, Japan
I. IGARASHI
Affiliation:
National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Obihiro, Hokkaido 080-8555, Japan
T. OYAMADA
Affiliation:
Department of Veterinary Parasitology, School of Veterinary Medicine and Animal Sciences, Kitasato University, Towada, Aomori 034-8628, Japan
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Abstract

We produced a mAb against the Babesia caballi extracellular merozoite termed mAb 2H2 and used it to screen a cDNA expression library prepared from B. caballi merozoite mRNA for highly expressed proteins. The complete nucleotide sequence of the cloned gene had 1547 nucleotides and contained a 36-nucleotide intron. The 1398 nucleotide open reading frame predicts a 51 kDa protein showing similarity to protein disulfide isomerase (PDI) from other species. The PDI gene had a predicted N-terminal signal sequence of 19 amino acids and a C-terminal tetrapeptide sequence (His-Thr-Glu-Leu; HTEL) for retention in lumen of the endoplasmic reticulum (ER). The recombinant protein expressed in baculovirus showed an apparent mass of 51 kDa, identical to that the native B. caballi protein. Moreover, the ER retention signal site (HTEL) of the recombinant protein retained its function in ER of insect cells. This 51 kDa protein was strongly expressed by extracelluar B. caballi merozoites in indirect immunofluorescence antibody tests, and was not expressed in the early phase of trophozoite development. Interestingly, detailed observation showed that the reaction of anti-P51 antibody and mAb 2H2 against pear-shaped forms was very erratic, some displaying one or two brightly fluorescent patterns.

Keywords

Babesia caballiprotein disulfide isomerasecDNAmAbindirect immunofluorescence antibody testendoplasmic reticulum
Type
Research Article
Information
Parasitology , Volume 131 , Issue 6 , December 2005 , pp. 775 - 782
Copyright
© 2005 Cambridge University Press

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References

REFERENCES

Avarazed, A., Igarashi, I., Kanemaru, K., Hirumi, K., Omata, Y., Saito, A., Oyamada, T., Nagasawa, H., Toyoda, Y. and Suzuki, N. ( 1997). Improved in vitro cultivation of Babesia caballi. Journal of Veterinary Medical Science 59, 479–481.CrossRefGoogle Scholar
Blackman, M. J. and Bannister, L. H. ( 2001). Apical organelles of Apicomplexa: biology and isolation by subcellular fractionation. Molecular and Biochemical Parasitology 117, 11–25.CrossRefGoogle Scholar
Brüning, A. ( 1996). Equine piroplasmosis: an update on diagnosis, treatment and prevention. British Veterinary Journal 152, 139–151.CrossRefGoogle Scholar
Carruthers, V. B. ( 1999). Armed and dangerous: Toxoplasma gondii uses an arsenal of secretory proteins to infect host cells. Parasitology International 48, 1–10.Google Scholar
Dubremetz, J. E., Garcia-Reguet, N., Conseil, V. and Fourmaux, M. N. ( 1998). Apical organelles and host-cell invasion by Apicomplexa. International Journal for Parasitology 28, 1007–1013.CrossRefGoogle Scholar
Feinberg, A. P. and Vogelstein, B. ( 1983). A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. Analytical Biochemistry 132, 6–13.CrossRefGoogle Scholar
Ferrari, D. M. and Soling, H. D. ( 1999). The protein disulfide-isomerase family: unravelling a string of folds. Biometrical Journal 339, 1–10.Google Scholar
Florent, I., Mouray, E., Dali Ali, F., Drobecq, H., Girault, S., Schrevel, J., Sergheraert, C. and Grellier, P. ( 2000). Cloning of Plasmodium falciparum protein disulfide isomerase homologue by affinity purification using the antiplasmodial inhibitor 1,4-bis[3-[N-(cyclohexyl methyl)amino]propyl]piperazine. FEBS Letters 484, 246–252.CrossRefGoogle Scholar
Ikadai, H., Martin, M. D., Nagasawa, H., Fujisaki, K., Suzuki, N., Mikami, T., Kudo, N., Oyamada, T. and Igarashi, I. ( 2001). Analysis of a growth-promoting factor for Babesia caballi cultivation. Journal of Parasitology 87, 1484–1486.CrossRefGoogle Scholar
Ikadai, H., Tamaki, Y., Xuan, X., Igarashi, I., Kawai, S., Nagasawa, H., Fujisaki, K., Toyoda, Y., Suzuki, N. and Mikami, T. ( 1999 a). Inhibitory effect of monoclonal antibodies on the growth of Babesia caballi. International Journal for Parasitology 29, 1785–1791.Google Scholar
Ikadai, H., Xuan, X., Igarashi, I., Tanaka, S., Kanemaru, T., Nagasawa, H., Fujisaki, K., Suzuki, N. and Mikami, T. ( 1999 b). Cloning and expression of a 48-kilodalton Babesia caballi merozoite rhoptry protein and potential use of the recombinant antigen in an enzyme-linked immunosorbent assay. Journal of Clinical Microbiology 37, 3475–3480.Google Scholar
Knowles, R. C. ( 1998). Equine babesiosis: epidemiology, control and chemotherapy. Equine Veterinary Science 8, 61–64.Google Scholar
Lyles, M. M. and Gilbert, H. F. ( 1994). Mutations in the thioredoxin sites of protein disulfide isomerase reveal functional nonequivalence of the N- and C-terminal domains. Journal of Biological Chemistry 269, 30946–30952.Google Scholar
Preiser, P., Kaviratne, M., Khan, S., Bannister, L. and Jarra, W. ( 2000). The apical organelles of malaria merozoites: host cell selection, invasion, host immunity and immune evasion. Microbe and Infection 2, 1461–1477.CrossRefGoogle Scholar
Sambrook, J., Fritsch, E. F. and Maniatis, T. ( 1989). Molecular Cloning: a Laboratory Manual, 2nd Edn. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, USA.
Schein, E. ( 1985). Equine babesiosis. In Babesiosis of Domestic Animals and Man ( ed. Ristic, M.) pp. 197–208. CRC press, Boca Raton, Florida, USA.
Soldati, D., Dubremetz, J. F. and Lebrun, M. ( 2001). Microneme proteins: structural and functional requirements to promote adhesion and invasion by the apicomplexan parasite Toxoplasma gondii. International Journal for Parasitology 31, 1293–1302.CrossRefGoogle Scholar
Von Heijne, G. ( 1986). A new method for predicting signal sequence cleavage sites. Nucleic Acids Research 14, 4683–4690.CrossRefGoogle Scholar
Vuori, K., Myllyla, R., Pihlajaniemi, T. and Kivirikko, K. I. ( 1992). Expression and site-directed mutagenesis of human protein disulfide isomerase in Escherichia coli. This multifunctional polypeptide has two independently acting catalytic sites for the isomerase activity. Journal of Biological Chemistry 267, 7211–7214.Google Scholar