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The Immunoregulatory Mediator Macrophage Migration Inhibitory Factor (MIF) Catalyzes a Tautomerization Reaction

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Abstract

Background

Recent studies of melanin biosynthesis have uncovered an unusual enzymatic activity which converts the non-naturally occurring D-isomer of 2-carboxy-2,3-dihydroindole-5,6-quinone (dopachrome) into 5,6-dihydroxyindole-2-carboxylic acid (DHICA). The aim of the present investigation was to isolate and characterize the enzyme catalyzing this tautomerization reaction.

Materials and Methods

After we performed a tissue survey of D-dopachrome tautomerase activity, 10 bovine lenses were homogenized and used as a source of enzyme. A soluble fraction was obtained by high-speed centrifugation and subjected to successive FPLC chromatography on Phenyl-sepharose, Mono S cation-exchange, and Superdex gel-filtration. The isolated enzyme was electrophoresed, blotted onto PVDF membrane, and the N terminus analyzed by gas phase micro-sequencing.

Results

The protein catalyzing the conversion of D-dopachrome to DHICA was purified to homogeneity in 14% yield and showed a molecular weight of 12 kD when analyzed by SDS-PAGE. The first 27 amino acid residues of this protein were sequenced and found to be identical with those of bovine macrophage migration inhibitory factor (MIF). The catalytic activity of native MIF was confirmed by studies of purified recombinant human MIF, which showed the same tautomerase activity. While L-dopachrome was not a substrate for this reaction, the methyl esters of the L- and D-isomers were found to be better substrates for MIF than D-dopachrome.

Conclusions

MIF has been described recently to be an anterior pituitary hormone and to be released from immune cells stimulated by low concentrations of glucocorticoids. Once secreted, MIF acts to control, or counter-regulate, the immunosuppressive effects of glucocorticoids on the immune system. Although the tested substrate, D-dopachrome, does not occur naturally, the observation that MIF has tautomerase activity suggests that MIF may mediate its biological effects by an enzymatic reaction. These data also offer a potential approach for the design of small molecule pharmacological inhibitors of MIF that may modulate its potent immunoregulatory effects in vivo.

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References

  1. Bernhagen J, Calandra T, Mitchell RA, et al. (1993) MIF is a pituitary-derived cytokine that potentiates lethal endotoxaemia. Nature 365: 756–759.

    Article  CAS  Google Scholar 

  2. Calandra T, Bernhagen J, Mitchell RA, Bucala R. (1994) The macrophage is an important and previously unrecognized source of macrophage migration inhibitory factor. J. Exp. Med. 179: 1895–1902.

    Article  CAS  Google Scholar 

  3. Nishino T, Bernhagen J, Shiiki H, Calandra T, Dohi K, Bucala R. (in press) Localization of macrophage migration inhibitory factor (MIF) to secretory granules within the corticotrophic and thyrotrophic cells of the pituitary gland. Mol. Med.

  4. Calandra T, Bernhagen J, Metz CN, et al. (1995) Identification of MIF as a glucocorticoid-induced mediator and counter-regulator of glucocorticoid Inhibition of cytokine production. Nature 377: 68–71.

    Article  CAS  Google Scholar 

  5. Wistow GJ, Shaughnessy MP, Lee DC, Hodin J, Zelenka PS. (1993) A macrophage migration inhibitory factor is expressed in the differentiating cells of the eye lens. Proc. Natl. Acad. Sci. U.S.A. 90: 1272–1275.

    Article  CAS  Google Scholar 

  6. Galat A, Riviere S, Bouet F. (1993) Purification of macrophage migration inhibitory factor (MIF) from bovine brain cytosol. FEBS Lett. 319: 233–236.

    Article  CAS  Google Scholar 

  7. Sakai M, Nishihira J, Hibiya Y, Koyama Y, Nishi S. (1994) Glutathione binding rat liver 13k protein is the homologue of the macrophage migration inhibitory factor. Biochem. Mol. Biol. Int. 33: 439–446.

    CAS  PubMed  Google Scholar 

  8. Prota G. (1992) Melanins and Melanogenesis. Academic Press, San Diego.

    Google Scholar 

  9. Pawelek JM. (1991) After dopachrome? Pigment Cell Res. 4: 53–62.

    Article  CAS  Google Scholar 

  10. Pawelek JM. (1990) Dopachrome conversion factor functions as an isomerase. Biochem. Biophys. Res. Commun. 166: 1328–1333.

    Article  CAS  Google Scholar 

  11. Tsukamoto K, Jackson U, Urabe K, Montague PM, Hearing VJ. (1992) A second tyrosinase-related protein, TRP-2, is a melanogenic enzyme termed DOPAchrome tautomerase. EMBOJ. 11: 519–526.

    Article  CAS  Google Scholar 

  12. Winder AJ, Wittbjer A, Rosengren E, Rorsman H. (1993) The mouse brown (b) locus protein has dopachrome tautomerase activity and is located in lysosomes in transfected fibroblasts. J. Cell Sci. 106: 153–166.

    CAS  PubMed  Google Scholar 

  13. Odh G, Hindmith A, Rosengren A-M, Rosengren E, Rorsman H. (1993) Isolation of a new tautomerase monitored by the conversion of D-dopachrome to 5,6-dihydroxyindole. Biochem. Biophys. Res. Commun. 197: 619–624.

    Article  CAS  Google Scholar 

  14. Zhang M, Åman P, Grubb A, et al. (1995) Cloning and sequencing of a cDNA encoding rat D-dopachrome tautomerase. FEBS Lett. 373: 203–206.

    Article  CAS  Google Scholar 

  15. Bernhagen J, Mitchell RA, Calandra T, Voelter W, Cerami A, Bucala R. (1994) Purification, bioactivity and secondary structure analysis of mouse and human macrophage migration inhibitory factor (MIF). Biochemistry 33: 14144–14155.

    Article  CAS  Google Scholar 

  16. Bradford MM. (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 72: 248–254.

    Article  CAS  Google Scholar 

  17. Lowry OH, Rosebrough NJ, Farr AL, Randall RJ. (1951) Protein measurement with the Folin phenol reagent. J. Biol. Chem. 193:265–275.

    CAS  PubMed  Google Scholar 

  18. Laemmli UK. (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 277: 680–685.

    Article  Google Scholar 

  19. Matsudaira P. (1987) Sequence from picomole quantities of proteins electroblotted onto polyvinylidene difluoride membranes. J. Biol. Chem. 262: 10035–10038.

    CAS  PubMed  Google Scholar 

  20. Aroca P, Solano R, García-Borron JC, Lozano JA. (1991) Specificity of dopachrome tautomerase and inhibition by carboxylated indoles. Biochem. J. 277: 393–397.

    Article  CAS  Google Scholar 

  21. Shibahara S, Tomita Y, Sakakura T, Nager C, Chadhuri B, Müller R. (1986) Cloning and expression of cDNA encoding mouse tyrosinase. Nucleic Acids Res. 14: 2413–2427.

    Article  CAS  Google Scholar 

  22. Jackson JJ. (1988) A cDNA encoding tyrosinase-related protein maps to the brown locus in mouse. Proc. Nat. Acad. Sci. U.S.A. 85: 4392–4396.

    Article  CAS  Google Scholar 

  23. Jackson JJ, Chambers DM, Tsukamoto K, et al. (1992) A second tyrosinase-related protein, TRP-2, maps to and is mutated at the mouse slaty locus. EMBO J. 11: 527–535.

    Article  CAS  Google Scholar 

  24. Sugumaran M, Semensi V. (1991) Quinone methide as a new intermediate in eumelanin biosynthesis. J. Biol. Chem. 266: 6073–6078.

    CAS  PubMed  Google Scholar 

  25. Sherry B, Yarlett N, Strupp A, Cerami A. (1992) Identification of cyclophilin as a proinflammatory secretory product of lipopolysaccharide-activated macrophages. Proc. Natl. Acad. Sci U.S.A. 89: 3511–3515.

    Article  CAS  Google Scholar 

  26. Schreiber SL, Crabtree GR. (1992) The mechanism of action of cyclosporin A and FK506. Immunol. Today 13: 136–142.

    Article  CAS  Google Scholar 

Download references

Acknowledgments

This work was supported by the Swedish Cancer Society; the Walter, Ellen, and Lennart Hesselman Foundation for Scientific Research; the Edvard Welander Foundation for Scientific Research; the Thure Carlsson Foundation for Scientific Research; the Anna Lisa and Sven-Eric Lundgren Foundation for Scientific Research; the Alfred Österlund Foundation for Scientific Research; the Donation Funds of the University Hospital, Lund; and National Institute of Health Grant A135931 (RB).

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Authors and Affiliations

  1. Department of Pharmacology, University Hospital, Lund, Sweden

    Evald Rosengren, Lars Jacobsson & Gerd Odh

  2. The Picower Institute for Medical Research, Manhasset, New York, USA

    Richard Bucala

  3. Department of Clinical Genetics, University Hospital, Lund, Sweden

    Pierre Åman & Christine N. Metz

  4. Department of Dermatology, University Hospital, S-22185, Lund, Sweden

    Hans Rorsman

Authors
  1. Evald Rosengren
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  2. Richard Bucala
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  3. Pierre Åman
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  4. Lars Jacobsson
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  5. Gerd Odh
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  6. Christine N. Metz
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  7. Hans Rorsman
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Additional information

Contributed by S. Moncada on November 14, 1995.

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Rosengren, E., Bucala, R., Åman, P. et al. The Immunoregulatory Mediator Macrophage Migration Inhibitory Factor (MIF) Catalyzes a Tautomerization Reaction. Mol Med 2, 143–149 (1996). https://doi.org/10.1007/BF03402210

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  • DOI: https://doi.org/10.1007/BF03402210

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