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. 1991 Apr 1;173(4):961–969. doi: 10.1084/jem.173.4.961

Hemoglobin degradation in the human malaria pathogen Plasmodium falciparum: a catabolic pathway initiated by a specific aspartic protease

PMCID: PMC2190804  PMID: 2007860

Abstract

Hemoglobin is an important nutrient source for intraerythrocytic malaria organisms. Its catabolism occurs in an acidic digestive vacuole. Our previous studies suggested that an aspartic protease plays a key role in the degradative process. We have now isolated this enzyme and defined its role in the hemoglobinolytic pathway. Laser desorption mass spectrometry was used to analyze the proteolytic action of the purified protease. The enzyme has a remarkably stringent specificity towards native hemoglobin, making a single cleavage between alpha 33Phe and 34Leu. This scission is in the hemoglobin hinge region, unraveling the molecule and exposing other sites for proteolysis. The protease is inhibited by pepstatin and has NH2-terminal homology to mammalian aspartic proteases. Isolated digestive vacuoles make a pepstatin- inhibitable cleavage identical to that of the purified enzyme. The pivotal role of this aspartic hemoglobinase in initiating hemoglobin degradation in the malaria parasite digestive vacuoles is demonstrated.

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Selected References

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  1. Aikawa M., Hepler P. K., Huff C. G., Sprinz H. The feeding mechanism of avian malarial parasites. J Cell Biol. 1966 Feb;28(2):355–373. doi: 10.1083/jcb.28.2.355. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Aissi E., Charet P., Bouquelet S., Biguet J. Endoprotease in Plasmodium yoelii nigeriensis. Comp Biochem Physiol B. 1983;74(3):559–566. doi: 10.1016/0305-0491(83)90229-8. [DOI] [PubMed] [Google Scholar]
  3. Azuma T., Pals G., Mohandas T. K., Couvreur J. M., Taggart R. T. Human gastric cathepsin E. Predicted sequence, localization to chromosome 1, and sequence homology with other aspartic proteinases. J Biol Chem. 1989 Oct 5;264(28):16748–16753. [PubMed] [Google Scholar]
  4. Baldwin J., Chothia C. Haemoglobin: the structural changes related to ligand binding and its allosteric mechanism. J Mol Biol. 1979 Apr 5;129(2):175–220. doi: 10.1016/0022-2836(79)90277-8. [DOI] [PubMed] [Google Scholar]
  5. Beavis R. C., Chait B. T. Cinnamic acid derivatives as matrices for ultraviolet laser desorption mass spectrometry of proteins. Rapid Commun Mass Spectrom. 1989 Dec;3(12):432–435. doi: 10.1002/rcm.1290031207. [DOI] [PubMed] [Google Scholar]
  6. Beavis R. C., Chait B. T. High-accuracy molecular mass determination of proteins using matrix-assisted laser desorption mass spectrometry. Anal Chem. 1990 Sep 1;62(17):1836–1840. doi: 10.1021/ac00216a020. [DOI] [PubMed] [Google Scholar]
  7. Beavis R. C., Chait B. T. Matrix-assisted laser-desorption mass spectrometry using 355 nm radiation. Rapid Commun Mass Spectrom. 1989 Dec;3(12):436–439. doi: 10.1002/rcm.1290031208. [DOI] [PubMed] [Google Scholar]
  8. Beavis R. C., Chait B. T. Rapid, sensitive analysis of protein mixtures by mass spectrometry. Proc Natl Acad Sci U S A. 1990 Sep;87(17):6873–6877. doi: 10.1073/pnas.87.17.6873. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Beychok S., Tyuma I., Benesch R. E., Benesch R. Optically active absorption bands of hemoglobin and its subunits. J Biol Chem. 1967 May 25;242(10):2460–2462. [PubMed] [Google Scholar]
  10. COOK L., GRANT P. T., KERMACK W. O. Proteolytic enzymes of the erythrocytic forms of roden and simian species of malarial plasmodia. Exp Parasitol. 1961 Nov;11:372–379. doi: 10.1016/0014-4894(61)90041-8. [DOI] [PubMed] [Google Scholar]
  11. Fagan J. M., Waxman L., Goldberg A. L. Red blood cells contain a pathway for the degradation of oxidant-damaged hemoglobin that does not require ATP or ubiquitin. J Biol Chem. 1986 May 5;261(13):5705–5713. [PubMed] [Google Scholar]
  12. Fitch C. D., Chevli R., Banyal H. S., Phillips G., Pfaller M. A., Krogstad D. J. Lysis of Plasmodium falciparum by ferriprotoporphyrin IX and a chloroquine-ferriprotoporphyrin IX complex. Antimicrob Agents Chemother. 1982 May;21(5):819–822. doi: 10.1128/aac.21.5.819. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. GROMAN N. B. Dynamic aspects of the nitrogen metabolism of Plasmodium gallinaceum in vivo and in vitro. J Infect Dis. 1951 Mar-Apr;88(2):126–150. doi: 10.1093/infdis/88.2.126. [DOI] [PubMed] [Google Scholar]
  14. Goldberg D. E., Slater A. F., Cerami A., Henderson G. B. Hemoglobin degradation in the malaria parasite Plasmodium falciparum: an ordered process in a unique organelle. Proc Natl Acad Sci U S A. 1990 Apr;87(8):2931–2935. doi: 10.1073/pnas.87.8.2931. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Gyang F. N., Poole B., Trager W. Peptidases from Plasmodium falciparum cultured in vitro. Mol Biochem Parasitol. 1982 Apr;5(4):263–273. doi: 10.1016/0166-6851(82)90034-2. [DOI] [PubMed] [Google Scholar]
  16. Hempelmann E., Wilson R. J. Endopeptidases from Plasmodium knowlesi. Parasitology. 1980 Apr;80(2):323–330. doi: 10.1017/s0031182000000780. [DOI] [PubMed] [Google Scholar]
  17. Krogstad D. J., Schlesinger P. H., Gluzman I. Y. Antimalarials increase vesicle pH in Plasmodium falciparum. J Cell Biol. 1985 Dec;101(6):2302–2309. doi: 10.1083/jcb.101.6.2302. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  19. Lambros C., Vanderberg J. P. Synchronization of Plasmodium falciparum erythrocytic stages in culture. J Parasitol. 1979 Jun;65(3):418–420. [PubMed] [Google Scholar]
  20. Levy M. R., Chou S. C. Activity and some properties of an acid proteinase from normal and Plasmodium berghei-infected red cells. J Parasitol. 1973 Dec;59(6):1064–1070. [PubMed] [Google Scholar]
  21. Levy M. R., Siddiqui W. A., Chou S. C. Acid protease activity in Plasmodium falciparum and P. knowlesi and ghosts of their respective host red cells. Nature. 1974 Feb 22;247(5442):546–549. doi: 10.1038/247546a0. [DOI] [PubMed] [Google Scholar]
  22. MORRISON D. B., JESKEY H. A. Alterations in some constituents of the monkey erythrocyte infected with Plasmodium knowlesi as related to pigment formation. J Natl Malar Soc. 1948 Dec;7(4):259–264. [PubMed] [Google Scholar]
  23. Matsudaira P. Sequence from picomole quantities of proteins electroblotted onto polyvinylidene difluoride membranes. J Biol Chem. 1987 Jul 25;262(21):10035–10038. [PubMed] [Google Scholar]
  24. Polet H., Conrad M. E. Malaria: extracellular amino acid requirements for in vitro growth of erythrocytic forms of Plasmodium knowlesi. Proc Soc Exp Biol Med. 1968 Jan;127(1):251–253. doi: 10.3181/00379727-127-32666. [DOI] [PubMed] [Google Scholar]
  25. Rosenthal P. J., McKerrow J. H., Aikawa M., Nagasawa H., Leech J. H. A malarial cysteine proteinase is necessary for hemoglobin degradation by Plasmodium falciparum. J Clin Invest. 1988 Nov;82(5):1560–1566. doi: 10.1172/JCI113766. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Roth E. F., Jr, Brotman D. S., Vanderberg J. P., Schulman S. Malarial pigment-dependent error in the estimation of hemoglobin content in Plasmodium falciparum-infected red cells: implications for metabolic and biochemical studies of the erythrocytic phases of malaria. Am J Trop Med Hyg. 1986 Sep;35(5):906–911. doi: 10.4269/ajtmh.1986.35.906. [DOI] [PubMed] [Google Scholar]
  27. Rudzinska M. A., Trager W., Bray R. S. Pinocytotic uptake and the digestion of hemoglobin in malaria parasites. J Protozool. 1965 Nov;12(4):563–576. doi: 10.1111/j.1550-7408.1965.tb03256.x. [DOI] [PubMed] [Google Scholar]
  28. Sato K., Fukabori Y., Suzuki M. Plasmodium berghei: a study of globinolytic enzyme in erythrocytic parasite. Zentralbl Bakteriol Mikrobiol Hyg A. 1987 May;264(3-4):487–495. doi: 10.1016/s0176-6724(87)80072-x. [DOI] [PubMed] [Google Scholar]
  29. Sherman I. W. Amino acid metabolism and protein synthesis in malarial parasites. Bull World Health Organ. 1977;55(2-3):265–276. [PMC free article] [PubMed] [Google Scholar]
  30. Sherman I. W. Biochemistry of Plasmodium (malarial parasites). Microbiol Rev. 1979 Dec;43(4):453–495. doi: 10.1128/mr.43.4.453-495.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Sherman I. W., Tanigoshi L. Purification of Plasmodium lophurae cathepsin D and its effects on erythrocyte membrane proteins. Mol Biochem Parasitol. 1983 Jul;8(3):207–226. doi: 10.1016/0166-6851(83)90044-0. [DOI] [PubMed] [Google Scholar]
  32. Tang J., Wong R. N. Evolution in the structure and function of aspartic proteases. J Cell Biochem. 1987 Jan;33(1):53–63. doi: 10.1002/jcb.240330106. [DOI] [PubMed] [Google Scholar]
  33. Theakston R. D., Fletcher K. A., Maegraith B. G. The use of electron microscope autoradiography for examining the uptake and degradation of haemoglobin by Plasmodium berghei. Ann Trop Med Parasitol. 1970 Mar;64(1):63–71. doi: 10.1080/00034983.1970.11686664. [DOI] [PubMed] [Google Scholar]
  34. Trager W., Jensen J. B. Human malaria parasites in continuous culture. Science. 1976 Aug 20;193(4254):673–675. doi: 10.1126/science.781840. [DOI] [PubMed] [Google Scholar]
  35. Vander Jagt D. L., Hunsaker L. A., Campos N. M. Characterization of a hemoglobin-degrading, low molecular weight protease from Plasmodium falciparum. Mol Biochem Parasitol. 1986 Mar;18(3):389–400. doi: 10.1016/0166-6851(86)90095-2. [DOI] [PubMed] [Google Scholar]
  36. Vander Jagt D. L., Intress C., Heidrich J. E., Mrema J. E., Rieckmann K. H., Heidrich H. G. Marker enzymes of Plasmodium falciparum and human erythrocytes as indicators of parasite purity. J Parasitol. 1982 Dec;68(6):1068–1071. [PubMed] [Google Scholar]
  37. Weber I. T., Miller M., Jaskólski M., Leis J., Skalka A. M., Wlodawer A. Molecular modeling of the HIV-1 protease and its substrate binding site. Science. 1989 Feb 17;243(4893):928–931. doi: 10.1126/science.2537531. [DOI] [PubMed] [Google Scholar]
  38. Wray W., Boulikas T., Wray V. P., Hancock R. Silver staining of proteins in polyacrylamide gels. Anal Biochem. 1981 Nov 15;118(1):197–203. doi: 10.1016/0003-2697(81)90179-2. [DOI] [PubMed] [Google Scholar]
  39. Zarchin S., Krugliak M., Ginsburg H. Digestion of the host erythrocyte by malaria parasites is the primary target for quinoline-containing antimalarials. Biochem Pharmacol. 1986 Jul 15;35(14):2435–2442. doi: 10.1016/0006-2952(86)90473-9. [DOI] [PubMed] [Google Scholar]

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