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Studies on the active site of human leukocytic pyrogen

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Abstract

Leukocytic pyrogen, a polypeptide produced by phagocytic mononuclear cells, is thought to be the endogenous mediator of fever. In addition to its effects on thermoregulation, leukocytic pyrogen has been shown to induce synthesis of acute-phase proteins, increase lymphocyte blastogenesis to mitogens, and cause release of neutrophil-specific granule contents. Despite its important role in biologic responses, little is known concerning the structure-function relationship of the molecule. In the present studies several protein-modifying conditions were used in order to examine specific amino acid participation at the active site. Because the state of purity of leukocytic pyrogen may be critical during certain reaction conditions, highly purified preparations were used. Experiments suggest that the active site requires the gamma-carboxyl group of glutamic acid and that blocking arginine reduces both the pyrogenic and neutrophil releasing properties of the molecule. Other studies demonstrate that the pyrogenicity of human leukocytic pyrogen is not due to serine esterase or carboxypeptidase B activity and that the 15,000-dalton molecule may be a glycoprotein. These experiments provide further evidence that the lymphocyte-activating and neutrophil-granule-releasing properties of human leukocytic pyrogen require the same active site which produces fever.

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References

  1. Kozak, M. S., H. H. Hahn, W. J. Lennarz, andW. B. Wood, Jr. 1968. Studies on the pathogenesis of fever. XVI. Purification and further chemical characterization of granulocytic pyrogen.J. Exp. Med. 127:341–357.

    Google Scholar 

  2. Bodel, P., A. Wechsler, andE. Atkins. 1969. Comparison of endogenous pyrogens from human and rabbit leukocytes utilizing Sephadex filtration.Yale J. Biol. Med. 41:376–387.

    Google Scholar 

  3. Moore, D. M., P. A. Murphy, J. P. Chesney, andW. B. Wood, Jr. 1973. Synthesis of endogenous pyrogen by rabbit leukocytes.J. Exp. Med. 137:1263–1274.

    Google Scholar 

  4. Dinarello, C. A., N. P. Goldin, andS. M. Wolff. 1974. Demonstration and characterization of two distinct human leukocytic pyrogens.J. Exp. Med. 139:1369–1381.

    Google Scholar 

  5. Murphy, P. A., J. P. Chesney, andW. B. Wood, Jr. 1974. Further purification of rabbit leukocyte pyrogen.J. Lab. Clin. Med. 83:310–322.

    Google Scholar 

  6. Dinarello, C. A., andS. M. Wolff. 1977. Partial purification of human leukocytic pyrogen.Inflammation 2:179–189.

    Google Scholar 

  7. Murphy, P. A., P. L. Simon, andW. F. Willoughby. 1980. Endogenous pyrogens made by rabbit peritoneal exudate cells are identical with lymphocyte-activating factors made by rabbit alveolar macrophages.J. Immunol. 124:2498–2501.

    Google Scholar 

  8. Cebula, T. A., D. F. Hansom, D. M. Moore, andP. A. Murphy. 1979. Synthesis of four endogenous pyrogens by rabbit macrophages.J. Lab. Clin. Med. 94:95–105.

    Google Scholar 

  9. Dinarello, C. A. 1980. Endogenous pyrogen.In Fever. J. M. Lipton, editor. Raven Press, New York. 1–9.

    Google Scholar 

  10. Bodel, P. 1974. Studies on the mechanism of endogenous pyrogen production. III. Human blood monocytes.J. Exp. Med. 140:954–965.

    Google Scholar 

  11. Hanson, D. F., P. A. Murphy, andB. E. Windle. 1980. Failure of rabbit neutrophils to secrete endogenous pyrogen when stimulated with staphylococci.J. Exp. Med. 151:1360–1371.

    Google Scholar 

  12. Nordlund, J. J., R. K. Root, andS. M. Wolff. 1970. Studies on the origin of human leukocytic pyrogen.J. Exp. Med. 131:727–743.

    Google Scholar 

  13. Dinarello, C. A. 1979. Production of endogenous pyrogen.Fed. Proc. 38:52–56.

    Google Scholar 

  14. Rosendorff, C., andJ. J. Mooney. 1971. Central nervous system sites of action of a purified leukocyte pyrogen.Am. J. Physiol. 220:597–603.

    Google Scholar 

  15. Lipton, J. M., andG. P. Trzcinka. 1976. Persistence of febrile response to pyrogens after PO/AH lesions in squirrel monkeys.Am. J. Physiol. 231:1638–1648.

    Google Scholar 

  16. Schoener, E. P., andS. C. Wang. 1975. Leukocytic pyrogen and sodium acetylsalicylate on hypothalamic neurons in the cat.Am. J. Physiol. 229:185–190.

    Google Scholar 

  17. Bernheim, H. A., T. M. Gelbiert, andJ. T. Stitt. 1980. Prostaglandin E levels in third ventricular cerebrospinal fluid of rabbits during fever and changes in body temperature.J. Physiol. 301:69–78.

    Google Scholar 

  18. Dinarello, C. A., andH. A. Bernheim. 1981. Ability of leukocytic pyrogen to stimulate brain prostaglandin in vitro.J. Neurochem. 37:702–708.

    Google Scholar 

  19. Klempner, M. S., C. A. Dinarello, andJ. I. Gallin. 1978. Human leukocytic pyrogen induces release of specific granule contents from human neutrophils.J. Clin. Invest. 61: 1330–1336.

    Google Scholar 

  20. Rosenwasser, L. J., C. A. Dinarello, andA. S. Rosenthal. 1979. Adherent cell function in murine T-lymphocyte antigen recognition. IV. Enhancement of murine T-cell antigen recognition by human leukocytic pyrogen.J. Exp. Med. 150:709–714.

    Google Scholar 

  21. McAdam, K. P. W. J., andC. A. Dinarello. 1980. Induction of serum amyloid A synthesis by human leukocytic pyrogen.In Bacterial Endotoxins and Host Response. M. K. Agarwal, editor. Elsvier-North Holland, Amsterdam. 167–178.

    Google Scholar 

  22. Hooper, D. C., C. J. Steer, C. A. Dinarello, andA. C. Peacock. 1981. Haptoglobin and albumin synthesis in isolated rat hepatocytes. Response to potential mediators of the acute-phase reaction.Biochim. Biophys. Acta 653:118–129.

    Google Scholar 

  23. Gordon, A. H., andI. D. Parker. 1980. A pyrogen from human cells which is active in mice.Br. J. Exp. Pathol 61:534–539.

    Google Scholar 

  24. Wolff, S. M., J. H. Mulholland, andS. B. Ward. 1965. Quantitative aspects of the pyrogenic responses of rabbits to endotoxin.J. Lab. Clin. Med. 65:268–276.

    Google Scholar 

  25. Elin, R. J., andS. M. Wolff. 1973. Nonspecificity of the Limulus amebocyte lysate test: positive reactions with polynucleotides and proteins.J. Infect. Dis. 128:349–352.

    Google Scholar 

  26. Root, R. K., J. J. Nordlund, andS. M. Wolff. 1970. Factors affecting the quantitative production and assay of human leukocytic pyrogen.J. Lab. Clin. Med. 75:679–693.

    Google Scholar 

  27. Dinarello, C. A., L. Renfer, andS. M. Wolff. 1977. The production of antibody against human leukocytic pyrogen.J. Clin. Invest. 60:465–472.

    Google Scholar 

  28. Dinarello, C. A., L. Renfer, andS. M. Wolff. 1977. Human leukocytic pyrogen: Purification and development of a radioimmunoassay.Proc. Natl. Acad. Sci. U.S.A. 74:4624–4627.

    Google Scholar 

  29. Petra, P. 1971. Modification of carboxyl groups in bovine carboxypeptidase. A. I. Inactivation of the enzyme by N-ethyl-5-phenylisoxazolium-3′-sulfonate (Woodward's reagent K).Biochemistry 10:3163–3170.

    Google Scholar 

  30. Marcus, F., S. M. Schuster, andH. A. Lardy. 1976. Essential arginyl residues in mitochondrial adenosine triphosphate.J. Biol. Chem. 254:1775–1780.

    Google Scholar 

  31. Takahashi, K. 1968. The reaction of phenylglyoxal with arginine residues in proteins.J. Biol. Chem. 243:6171–6179.

    Google Scholar 

  32. Takahashi, K. 1977. The reactions of phenylglyoxal and related reagents with amino acids.J. Biochem. 81:395–402.

    Google Scholar 

  33. Kobashi, K., andB. L. Horecker. 1967. Reversible inactivation of rabbit muscle aldolase byo-phenantroline.Arch. Biochem. Biophys. 121:178.

    Google Scholar 

  34. Steck, T. L. 1972. Cross-linking the major proteins of the isolated erythrocyte membrane.J. Mol Biol. 66:295–305.

    Google Scholar 

  35. Werber, M. M., andM. Sokolovsky. 1972. Chemical evidence for a functional arginine residue in carboxypeptidase B.Biochem. Biophys. Res. Commun. 48:384–390.

    Google Scholar 

  36. Plummer, T. H., Jr. 1969. Isolation and sequence of peptides at the active center of bovine carboxypeptidase B.J. Biol. Chem. 244:5246–5253.

    Google Scholar 

  37. Waksman, B. H., J.-P. Dessaint, andS. P. Katz. 1980. Proteolysis, calcium and cyclic nucleotides in macrophage T-lymphocyte interaction.In Biochemical Characterization of Lymphokines. A. L. de Weck, F. Kristensen, and M. Landy, editor. Acaademic Press, New York. 435–438.

    Google Scholar 

  38. Dinarello, C. A., andL. J. Rosenwasser. 1981. Lymphocyte activating property of human leukocytic pyrogen.In Advances in Immunopharmacology. J. Hadden, L. Chedid, P. Mullen, and F. Spreafico, editors. Pergammon Press, Oxford. 419–425.

    Google Scholar 

  39. Rossenwasser, L. J., andC. A. Dinarello. 1981. Ability of human leukocytic pyrogen to enhance phytohemagglutinin induced murine thymocyte proliferation.Cell. Immunol. 63: 134–142.

    Google Scholar 

  40. Sztein, M. B., S. N. Vogel, J. D. Sipe, P. A. Murphy, S. B. Mizel, J. J. Oppenheim, andD. L. Rosenstreich. 1981. The role of macrophages in the acute-phase response: SAA inducer is closely related to lymphocyte activating factor and endogenous pyrogen.Cell. Immunol. 63:164–176.

    Google Scholar 

  41. Plummer, T. H., Jr. 1971. Evidence for a carboxyl group at the active center of bovine carboxypeptidase B.J. Biol. Chem. 246:2930–2935.

    Google Scholar 

  42. Benutzen, K. 1977. Human leukocyte migration inhibitory factor (LIF). II. Partial biochemical characterization of the substrate specificities for this lymphokine.Scand. J. Immunol. 6:133–140.

    Google Scholar 

  43. Clamp, J. R., andHough, L. 1965. The periodate oxidation of amino acids with reference to studies on glycoproteins.Biochem. J. 94:17–22.

    Google Scholar 

  44. Lachman, L. B., andR. S. Metzoar. 1980. Purification and characterization of human lymphocyte activating factor.In Biochemical Characterization of Lymphokines. A. L. de Weck, F. Kristensen, and M. Landy, editors. Academic Press, New York. 405–409.

    Google Scholar 

  45. Dinarello, C. A. 1981. Quantitation of endogenous pyrogens.In Methods for Studying Mononuclear Phagocytes. D. O. Adams, H. Koren, and P. Edelson, editor. Academic Press, New York. 629–639.

    Google Scholar 

  46. Bernheim, H. A., andKluger, M. J. 1977. Endogenous pyrogen-like substance produced by reptiles.J. Physiol. 267:659–666.

    Google Scholar 

  47. Murphy, P. A., D. F. Hanson, P. L. Simon, W. F. Willoughby, andB. E. Windle. 1980. Properties of two distinct endogenous pyrogens secreted by rabbit macrophages.In Microbiology. D. Schlesinger, editor. American Society for Microbiology, Washington D.C. 158–161.

    Google Scholar 

  48. Sigman, D. S., andMooser, G. 1975. Chemical studies of enzyme active sites.Annu. Rev. Biochem. 44:889–931.

    Google Scholar 

  49. Clark, R. A., S. Szot, K. Venkatasubramanian, andE. Schiffman. 1980. Chemotactic factor inactivation by myeloperoxidase-mediated oxidation of methionine.J. Immunol. 124:2020–2026.

    Google Scholar 

  50. Kycia, J. H., M. Elzinga, N. Alonza, andC. H. W. Hins. 1968. Primary structure of bovine carboxypeptidase B.Arch. Biochem. Biophys. 123:336–342.

    Google Scholar 

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Authors and Affiliations

  1. Department of Medicine, Division of Experimental Medicine, Tufts University School of Medicine, 02111, Boston, Massachusetts

    Charles A. Dinarello, Klaus Bendtzen & Sheldon M. Wolff

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  1. Charles A. Dinarello
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  2. Klaus Bendtzen
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  3. Sheldon M. Wolff
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Dinarello, C.A., Bendtzen, K. & Wolff, S.M. Studies on the active site of human leukocytic pyrogen. Inflammation 6, 63–78 (1982). https://doi.org/10.1007/BF00910720

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