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. 2003 Jan 1;369(Pt 1):191–198. doi: 10.1042/BJ20021402

Echinococcus granulosus antigen 5 is closely related to proteases of the trypsin family.

Carmen Lorenzo 1, Gustavo Salinas 1, Andreina Brugnini 1, Christer Wernstedt 1, Ulf Hellman 1, Gualberto González-Sapienza 1
PMCID: PMC1223071  PMID: 12358601

Abstract

Antigen 5 (Ag5) is a dominant secreted component of the larval stage of Echinococcus granulosus, and is highly immunogenic in human infections. Although the diagnostic value of Ag5 has been thoroughly evaluated, there has been little progress in its molecular characterization and the understanding of its biological role. In the present study, the Ag5 gene was cloned by reverse transcription-PCR on the basis of the amino acid sequences of tryptic fragments. The nucleotide sequence indicates that Ag5 is synthesized as a single polypeptide chain that is afterwards processed into single disulphide-bridged 22 and 38 kDa subunits. Whereas the 22 kDa component contains a highly conserved glycosaminoglycan-binding motif that may help to confine Ag5 in the host tissue surrounding the parasite, the 38 kDa subunit is closely related to serine proteases of the trypsin family. The sequences in the vicinity of the active-site histidine, aspartic acid and serine residues, and critical cysteine residues involved in disulphide formation, are well conserved, but the catalytic serine residue is replaced by threonine. Since there are no significant chemical differences between the O gamma atoms of these residues, we performed a series of enzymic assays to find out whether Ag5 is a catalytic molecule. Neither proteolytic activity nor binding to protease inhibitors could be detected using the native purified antigen. Thus it may be possible that Ag5 possesses a highly specific physiological substrate or, more likely, that trypsin-like folding has been recruited to fulfil novel functions.

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Selected References

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  1. Barbieri M., Fernández V., González G., Luaces V. M., Nieto A. Diagnostic evaluation of a synthetic peptide derived from a novel antigen B subunit as related to other available peptides and native antigens used for serology of cystic hydatidosis. Parasite Immunol. 1998 Feb;20(2):51–61. doi: 10.1046/j.1365-3024.1998.00117.x. [DOI] [PubMed] [Google Scholar]
  2. Bout D., Fruit J., Capron A. Purification d'un antigène spécifique de liquide hydatique. Ann Immunol (Paris) 1974 Aug-Sep;125 100(5):775–788. [PubMed] [Google Scholar]
  3. Brehm K., Jensen K., Frosch M. mRNA trans-splicing in the human parasitic cestode Echinococcus multilocularis. J Biol Chem. 2000 Dec 8;275(49):38311–38318. doi: 10.1074/jbc.M006091200. [DOI] [PubMed] [Google Scholar]
  4. Di Felice G., Pini C., Afferni C., Vicari G. Purification and partial characterization of the major antigen of Echinococcus granulosus (antigen 5) with monoclonal antibodies. Mol Biochem Parasitol. 1986 Aug;20(2):133–142. doi: 10.1016/0166-6851(86)90025-3. [DOI] [PubMed] [Google Scholar]
  5. Facon B., Chamekh M., Dissous C., Capron A. Molecular cloning of an Echinococcus granulosus protein expressing an immunogenic epitope of antigen 5. Mol Biochem Parasitol. 1991 Apr;45(2):233–239. doi: 10.1016/0166-6851(91)90090-s. [DOI] [PubMed] [Google Scholar]
  6. Gantt S. M., Clavijo P., Bai X., Esko J. D., Sinnis P. Cell adhesion to a motif shared by the malaria circumsporozoite protein and thrombospondin is mediated by its glycosaminoglycan-binding region and not by CSVTCG. J Biol Chem. 1997 Aug 1;272(31):19205–19213. doi: 10.1074/jbc.272.31.19205. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Gonzalez G., Orn A., Cazzulo J. J., Grönvik K. O. Production and characterization of monoclonal antibodies against the major cysteine proteinase of Trypanosoma cruzi. Scand J Immunol. 1994 Oct;40(4):389–394. doi: 10.1111/j.1365-3083.1994.tb03479.x. [DOI] [PubMed] [Google Scholar]
  8. González G., Nieto A., Fernández C., Orn A., Wernstedt C., Hellman U. Two different 8 kDa monomers are involved in the oligomeric organization of the native Echinococcus granulosus antigen B. Parasite Immunol. 1996 Dec;18(12):587–596. doi: 10.1046/j.1365-3024.1996.d01-38.x. [DOI] [PubMed] [Google Scholar]
  9. González G., Spinelli P., Lorenzo C., Hellman U., Nieto A., Willis A., Salinas G. Molecular characterization of P-29, a metacestode-specific component of Echinococcus granulosus which is immunologically related to, but distinct from, antigen 5. Mol Biochem Parasitol. 2000 Feb 5;105(2):177–184. doi: 10.1016/s0166-6851(99)00166-8. [DOI] [PubMed] [Google Scholar]
  10. Gottstein B., Witassek F., Eckert J. Neues zur Echinokokkose. Schweiz Med Wochenschr. 1986 Jun 14;116(24):810–817. [PubMed] [Google Scholar]
  11. Hellman U., Wernstedt C., Góez J., Heldin C. H. Improvement of an "In-Gel" digestion procedure for the micropreparation of internal protein fragments for amino acid sequencing. Anal Biochem. 1995 Jan 1;224(1):451–455. doi: 10.1006/abio.1995.1070. [DOI] [PubMed] [Google Scholar]
  12. Ichinose A., Takeya H., Espling E., Iwanaga S., Kisiel W., Davie E. W. Amino acid sequence of human protein Z, a vitamin K-dependent plasma glycoprotein. Biochem Biophys Res Commun. 1990 Nov 15;172(3):1139–1144. doi: 10.1016/0006-291x(90)91566-b. [DOI] [PubMed] [Google Scholar]
  13. Lesk A. M., Fordham W. D. Conservation and variability in the structures of serine proteinases of the chymotrypsin family. J Mol Biol. 1996 May 10;258(3):501–537. doi: 10.1006/jmbi.1996.0264. [DOI] [PubMed] [Google Scholar]
  14. Lightowlers M. W., Liu D. Y., Haralambous A., Rickard M. D. Subunit composition and specificity of the major cyst fluid antigens of Echinococcus granulosus. Mol Biochem Parasitol. 1989 Dec;37(2):171–182. doi: 10.1016/0166-6851(89)90149-7. [DOI] [PubMed] [Google Scholar]
  15. March F., Enrich C., Mercader M., Sánchez F., Muñoz C., Coll P., Prats G. Echinococcus granulosus: antigen characterization by chemical treatment and enzymatic deglycosylation. Exp Parasitol. 1991 Nov;73(4):433–439. doi: 10.1016/0014-4894(91)90067-7. [DOI] [PubMed] [Google Scholar]
  16. Morgan J. G., Sukiennicki T., Pereira H. A., Spitznagel J. K., Guerra M. E., Larrick J. W. Cloning of the cDNA for the serine protease homolog CAP37/azurocidin, a microbicidal and chemotactic protein from human granulocytes. J Immunol. 1991 Nov 1;147(9):3210–3214. [PubMed] [Google Scholar]
  17. Musiani P., Piantelli M., Lauriola L., Arru E., Pozzuoli R. Echinococcus granulosus: specific quantification of the two most immunoreactive antigens in hydatid fluids. J Clin Pathol. 1978 May;31(5):475–478. doi: 10.1136/jcp.31.5.475. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Oinonen C., Tikkanen R., Rouvinen J., Peltonen L. Three-dimensional structure of human lysosomal aspartylglucosaminidase. Nat Struct Biol. 1995 Dec;2(12):1102–1108. doi: 10.1038/nsb1295-1102. [DOI] [PubMed] [Google Scholar]
  19. Palm G. J., Lubkowski J., Derst C., Schleper S., Röhm K. H., Wlodawer A. A covalently bound catalytic intermediate in Escherichia coli asparaginase: crystal structure of a Thr-89-Val mutant. FEBS Lett. 1996 Jul 22;390(2):211–216. doi: 10.1016/0014-5793(96)00660-6. [DOI] [PubMed] [Google Scholar]
  20. Pozzuoli R., Musiani P., Arru E., Patrono C., Piantelli M. Echinococcus granulosus: evaluation of purified antigens immunoreactivity. Exp Parasitol. 1974 Feb;35(1):52–60. doi: 10.1016/0014-4894(74)90006-x. [DOI] [PubMed] [Google Scholar]
  21. Rost B., Sander C. Prediction of protein secondary structure at better than 70% accuracy. J Mol Biol. 1993 Jul 20;232(2):584–599. doi: 10.1006/jmbi.1993.1413. [DOI] [PubMed] [Google Scholar]
  22. Seemüller E., Lupas A., Stock D., Löwe J., Huber R., Baumeister W. Proteasome from Thermoplasma acidophilum: a threonine protease. Science. 1995 Apr 28;268(5210):579–582. doi: 10.1126/science.7725107. [DOI] [PubMed] [Google Scholar]
  23. Shepherd J. C., McManus D. P. Specific and cross-reactive antigens of Echinococcus granulosus hydatid cyst fluid. Mol Biochem Parasitol. 1987 Sep;25(2):143–154. doi: 10.1016/0166-6851(87)90003-x. [DOI] [PubMed] [Google Scholar]
  24. Siles-Lucas M., Gottstein B., Felleisen R. S. Identification of a differentially expressed Echinococcus multilocularis protein Em6 potentially related to antigen 5 of Echinococcus granulosus. Parasite Immunol. 1998 Oct;20(10):473–481. doi: 10.1046/j.1365-3024.1998.00178.x. [DOI] [PubMed] [Google Scholar]
  25. Wassell J. Haptoglobin: function and polymorphism. Clin Lab. 2000;46(11-12):547–552. [PubMed] [Google Scholar]
  26. Willis A. C., Díaz A., Nieto A., Sim R. B. Echinococcus granulosus antigen 5 may be a serine proteinase. Parasite Immunol. 1997 Aug;19(8):385–385. doi: 10.1046/j.1365-3024.1997.d01-222.x. [DOI] [PubMed] [Google Scholar]
  27. Zhang L. H., McManus D. P. Purification and N-terminal amino acid sequencing of Echinococcus granulosus antigen 5. Parasite Immunol. 1996 Dec;18(12):597–606. doi: 10.1046/j.1365-3024.1996.d01-42.x. [DOI] [PubMed] [Google Scholar]

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