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. 1985 May 1;161(5):1196–1212. doi: 10.1084/jem.161.5.1196

gp72, the 72 kilodalton glycoprotein, is the membrane acceptor site for C3 on Trypanosoma cruzi epimastigotes

PMCID: PMC2187612  PMID: 3886827

Abstract

We examined the interaction of complement component C3 with surface molecules on Trypanosoma cruzi. Five- to six-fold more C3 was bound to epimastigotes (Epi) than to metacyclic trypomastigotes (CMT) of strain M88. Epi and CMT were surface iodinated, then incubated in C8-deficient serum, and detergent lysates were applied to anti-C3 antibody that had been coupled to Sepharose. We found that 9.20-10.24% of applied 125I- Epi protein bound to anti-C3-sepharose, compared to 2.64% binding of 125I-CMT protein. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed that C3 was attached to 125I-Epi protein by a covalent bond. Samples eluted from anti-C3-sepharose with hydroxylamine revealed a single, major, 72 kD band, suggesting that C3b attaches almost exclusively to the 72 kD glycoprotein of Epi by a hydroxylamine- susceptible ester bond. An antiserum was prepared from lysates of serum- treated Epi that had been affinity-purified on anti-C3-sepharose. This antiserum immunoprecipitated a single 72 kD component (gp72) from surface-iodinated Epi, and specifically recognized only gp72 from Epi in immunoblots. In contrast to the results with Epi, gp72 on CMT was not found to be an efficient acceptor molecule for C3 deposition. The results are the first to evaluate the acceptor site for C3 deposition on a parasite, and they show that gp72 on Epi, but not gp72 on CMT, serves as the preferential acceptor for C3 during antibody-independent alternative complement pathway activation.

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Selected References

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  1. Alves M. J., Colli W. Glycoproteins from trypanosoma cruzi: partial purification by gel chromatography. FEBS Lett. 1975 Apr 1;52(2):188–190. doi: 10.1016/0014-5793(75)80803-9. [DOI] [PubMed] [Google Scholar]
  2. Alves M. J., Da Silveira J. F., De Paiva C. H., Tanaka C. T., Colli W. Evidence for the plasma membrane localization of carbohydrate-containing macromolecules from epimastigote forms of Trypanosoma cruzi. FEBS Lett. 1979 Mar 1;99(1):81–85. doi: 10.1016/0014-5793(79)80254-9. [DOI] [PubMed] [Google Scholar]
  3. Andrews N. W., Katzin A. M., Colli W. Mapping of surface glycoproteins of Trypanosoma cruzi by two-dimensional electrophoresis. A correlation with the cell invasion capacity. Eur J Biochem. 1984 May 2;140(3):599–604. doi: 10.1111/j.1432-1033.1984.tb08144.x. [DOI] [PubMed] [Google Scholar]
  4. Araujo F. G., Remington J. S. Characterization of stages and strains of Trypanosoma cruzi by analysis of cell membrane components. J Immunol. 1981 Sep;127(3):855–859. [PubMed] [Google Scholar]
  5. Brown E. J., Berger M., Joiner K. A., Frank M. M. Classical complement pathway activation by antipneumococcal antibodies leads to covalent binding of C3b to antibody molecules. Infect Immun. 1983 Nov;42(2):594–598. doi: 10.1128/iai.42.2.594-598.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Daha M. R., Fearon D. T., Austen K. F. C3 requirements for formation of alternative pathway C5 convertase. J Immunol. 1976 Aug;117(2):630–634. [PubMed] [Google Scholar]
  7. De Lederkremer R. M., Alves M. J., Fonseca G. C., Colli W. A lipopeptidophosphoglycan from Trypanosoma cruzi (epimastigota). Isolation, purification and carbohydrate composition. Biochim Biophys Acta. 1976 Aug 24;444(1):85–96. doi: 10.1016/0304-4165(76)90226-9. [DOI] [PubMed] [Google Scholar]
  8. Edwards M. S., Nicholson-Weller A., Baker C. J., Kasper D. L. The role of specific antibody in alternative complement pathway-mediated opsonophagocytosis of type III, group B Streptococcus. J Exp Med. 1980 May 1;151(5):1275–1287. doi: 10.1084/jem.151.5.1275. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Engel J. C., Dvorak J. A., Segura E. L., Crane M. S. Trypanosoma cruzi: biological characterization of 19 clones derived from two chronic chagasic patients. I. Growth kinetics in liquid medium. J Protozool. 1982 Nov;29(4):555–560. doi: 10.1111/j.1550-7408.1982.tb01334.x. [DOI] [PubMed] [Google Scholar]
  10. Fearon D. T. Regulation by membrane sialic acid of beta1H-dependent decay-dissociation of amplification C3 convertase of the alternative complement pathway. Proc Natl Acad Sci U S A. 1978 Apr;75(4):1971–1975. doi: 10.1073/pnas.75.4.1971. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Ferguson M. A., Allen A. K., Snary D. Studies on the structure of a phosphoglycoprotein from the parasitic protozoan Trypanosoma cruzi. Biochem J. 1983 Aug 1;213(2):313–319. doi: 10.1042/bj2130313. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Gadd K. J., Reid K. B. The binding of complement component C3 to antibody-antigen aggregates after activation of the alternative pathway in human serum. Biochem J. 1981 May 1;195(2):471–480. doi: 10.1042/bj1950471. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Gottlieb M. A carbohydrate-containing antigen from Trypanosoma cruzi and its detection in the circulation of infected mice. J Immunol. 1977 Aug;119(2):465–470. [PubMed] [Google Scholar]
  14. Hammer C. H., Wirtz G. H., Renfer L., Gresham H. D., Tack B. F. Large scale isolation of functionally active components of the human complement system. J Biol Chem. 1981 Apr 25;256(8):3995–4006. [PubMed] [Google Scholar]
  15. Howard R. J., Kaushal D. C., Carter R. Radioiodination of parasite antigens with 1,3,4,6-tetrachloro-3 alpha, 6 alpha-diphenylglycoluril (IODOGEN): studies with zygotes of Plasmodium gallinaceum. J Protozool. 1982 Feb;29(1):114–117. doi: 10.1111/j.1550-7408.1982.tb02891.x. [DOI] [PubMed] [Google Scholar]
  16. Joiner K. A., Goldman R., Schmetz M., Berger M., Hammer C. H., Frank M. M., Leive L. A quantitative analysis of C3 binding to O-antigen capsule, lipopolysaccharide, and outer membrane protein of E. coli 0111B4. J Immunol. 1984 Jan;132(1):369–375. [PubMed] [Google Scholar]
  17. Joiner K. A., Hammer C. H., Brown E. J., Cole R. J., Frank M. M. Studies on the mechanism of bacterial resistance to complement-mediated killing. I. Terminal complement components are deposited and released from Salmonella minnesota S218 without causing bacterial death. J Exp Med. 1982 Mar 1;155(3):797–808. doi: 10.1084/jem.155.3.797. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Kipnis T. L., David J. R., Alper C. A., Sher A., da Silva W. D. Enzymatic treatment transforms trypomastigotes of Trypanosoma cruzi into activators of alternative complement pathway and potentiates their uptake by macrophages. Proc Natl Acad Sci U S A. 1981 Jan;78(1):602–605. doi: 10.1073/pnas.78.1.602. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Kirchhoff L. V., Hieny S., Shiver G. M., Snary D., Sher A. Cryptic epitope explains the failure of a monoclonal antibody to bind to certain isolates of Trypanosoma cruzi. J Immunol. 1984 Nov;133(5):2731–2735. [PubMed] [Google Scholar]
  20. Lanham S. M., Godfrey D. G. Isolation of salivarian trypanosomes from man and other mammals using DEAE-cellulose. Exp Parasitol. 1970 Dec;28(3):521–534. doi: 10.1016/0014-4894(70)90120-7. [DOI] [PubMed] [Google Scholar]
  21. Law S. K., Levine R. P. Interaction between the third complement protein and cell surface macromolecules. Proc Natl Acad Sci U S A. 1977 Jul;74(7):2701–2705. doi: 10.1073/pnas.74.7.2701. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Law S. K., Lichtenberg N. A., Levine R. P. Evidence for an ester linkage between the labile binding site of C3b and receptive surfaces. J Immunol. 1979 Sep;123(3):1388–1394. [PubMed] [Google Scholar]
  23. Law S. K., Minich T. M., Levine R. P. Binding reaction between the third human complement protein and small molecules. Biochemistry. 1981 Dec 22;20(26):7457–7463. doi: 10.1021/bi00529a020. [DOI] [PubMed] [Google Scholar]
  24. Mann J., O'Brien R., Hostetter M. K., Alper C. A., Rosen F. S., Babior B. M. The third component of complement: covalent attachment of a radioactive sugar to the labile binding site of C3 via the alternative pathway. J Immunol. 1981 Jun;126(6):2370–2372. [PubMed] [Google Scholar]
  25. Markwell M. A., Haas S. M., Bieber L. L., Tolbert N. E. A modification of the Lowry procedure to simplify protein determination in membrane and lipoprotein samples. Anal Biochem. 1978 Jun 15;87(1):206–210. doi: 10.1016/0003-2697(78)90586-9. [DOI] [PubMed] [Google Scholar]
  26. Medicus R. G., Götze O., Müller-Eberhard H. J. Alternative pathway of complement: recruitment of precursor properdin by the labile C3/C5 convertase and the potentiation of the pathway. J Exp Med. 1976 Oct 1;144(4):1076–1093. doi: 10.1084/jem.144.4.1076. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Nogueira N., Bianco C., Cohn Z. Studies on the selective lysis and purification of Trypanosoma cruzi. J Exp Med. 1975 Jul 1;142(1):224–229. doi: 10.1084/jem.142.1.224. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Nogueira N., Chaplan S., Tydings J. D., Unkeless J., Cohn Z. Trypanosoma cruzi. Surface antigens of blood and culture forms. J Exp Med. 1981 Mar 1;153(3):629–639. doi: 10.1084/jem.153.3.629. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Nogueira N., Unkeless J., Cohn Z. Specific glycoprotein antigens on the surface of insect and mammalian stages of Trypanosoma cruzi. Proc Natl Acad Sci U S A. 1982 Feb;79(4):1259–1263. doi: 10.1073/pnas.79.4.1259. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Orellano E., Cazzulo J. J. Purification and regulatory properties of the NADP-linked malic enzyme for Crithidia fasciculata. Mol Biochem Parasitol. 1981 May;3(1):1–11. doi: 10.1016/0166-6851(81)90072-4. [DOI] [PubMed] [Google Scholar]
  31. Sher A., Crane M. S., Kirchhoff L. V. Incubation in mice provides a signal for the differentiation of Trypanosoma cruzi epimastigotes to trypomastigotes. J Protozool. 1983 May;30(2):278–283. doi: 10.1111/j.1550-7408.1983.tb02916.x. [DOI] [PubMed] [Google Scholar]
  32. Sher A., Snary D. Specific inhibition of the morphogenesis of Trypanosoma cruzi by a monoclonal antibody. Nature. 1982 Dec 16;300(5893):639–640. doi: 10.1038/300639a0. [DOI] [PubMed] [Google Scholar]
  33. Snary D. Cell surface glycoproteins of Trypanosoma cruzi: protective immunity in mice and antibody levels in human chagasic sera. Trans R Soc Trop Med Hyg. 1983;77(1):126–129. doi: 10.1016/0035-9203(83)90037-8. [DOI] [PubMed] [Google Scholar]
  34. Sullivan J. J. Metacyclogenesis of Trypanosoma cruzi in vitro: a simplified procedure. Trans R Soc Trop Med Hyg. 1982;76(3):300–303. doi: 10.1016/0035-9203(82)90173-0. [DOI] [PubMed] [Google Scholar]
  35. Tack B. F., Harrison R. A., Janatova J., Thomas M. L., Prahl J. W. Evidence for presence of an internal thiolester bond in third component of human complement. Proc Natl Acad Sci U S A. 1980 Oct;77(10):5764–5768. doi: 10.1073/pnas.77.10.5764. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Takata Y., Tamura N., Fujita T. Interaction of C3 with antigen-antibody complexes in the process of solubilization of immune precipitates. J Immunol. 1984 May;132(5):2531–2537. [PubMed] [Google Scholar]
  37. Venkatesh Y. P., Minich T. M., Law S. K., Levine R. P. Natural release of covalently bound C3b from cell surfaces and the study of this phenomenon in the fluid-phase system. J Immunol. 1984 Mar;132(3):1435–1439. [PubMed] [Google Scholar]

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