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. 1995 Aug 29;92(18):8383–8387. doi: 10.1073/pnas.92.18.8383

Leukotriene A4 hydrolase: mapping of a henicosapeptide involved in mechanism-based inactivation.

M J Mueller 1, A Wetterholm 1, M Blomster 1, H Jörnvall 1, B Samuelsson 1, J Z Haeggström 1
PMCID: PMC41161  PMID: 7667299

Abstract

Leukotriene A4 (LTA4) hydrolase [7E,9E,11Z,14Z)-(5S,6S)-5,6-epoxyicosa-7,9 ,11,14-tetraenoate hydrolase; EC 3.3.2.6] is a bifunctional zinc metalloenzyme which converts LTA4 into the chemotactic agent leukotriene B4 (LTB4). Suicide inactivation, a typical feature of LTA4 hydrolase/aminopeptidase, occurs via an irreversible, apparently mechanism-based, covalent binding of LTA4 to the protein in a 1:1 stoichiometry. Differential lysine-specific peptide mapping of unmodified and suicide-inactivated LTA4 hydrolase has been used to identify a henicosapeptide, encompassing the amino acid residues 365-385 of human LTA4 hydrolase, which is involved in the binding of LTA4, LTA4 methyl ester, and LTA4 ethyl ester to the native enzyme. A modified form of this peptide, generated by lysine-specific digestion of LTA4 hydrolase inactivated by LTA4 ethyl ester, could be isolated for complete Edman degradation. The sequence analysis revealed a gap at position 14, which shows that binding of the leukotriene epoxide had occurred via Tyr-378 in LTA4 hydrolase. Inactivation of the epoxide hydrolase and the aminopeptidase activity was accompanied by a proportionate modification of the peptide. Furthermore, both enzyme inactivation and peptide modification could be prevented by preincubation of LTA4 hydrolase with the competitive inhibitor bestatin, which demonstrates that the henicosapeptide contains functional elements of the active site(s). It may now be possible to clarify the molecular mechanisms underlying suicide inactivation and epoxide hydrolysis by site-directed mutagenesis combined with structural analysis of the lipid molecule, covalently bound to the peptide.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Arand M., Grant D. F., Beetham J. K., Friedberg T., Oesch F., Hammock B. D. Sequence similarity of mammalian epoxide hydrolases to the bacterial haloalkane dehalogenase and other related proteins. Implication for the potential catalytic mechanism of enzymatic epoxide hydrolysis. FEBS Lett. 1994 Feb 7;338(3):251–256. doi: 10.1016/0014-5793(94)80278-5. [DOI] [PubMed] [Google Scholar]
  2. Evans J. F., Kargman S. Bestatin inhibits covalent coupling of [3H]LTA4 to human leukocyte LTA4 hydrolase. FEBS Lett. 1992 Feb 3;297(1-2):139–142. doi: 10.1016/0014-5793(92)80345-h. [DOI] [PubMed] [Google Scholar]
  3. Evans J. F., Nathaniel D. J., Zamboni R. J., Ford-Hutchinson A. W. Leukotriene A3. A poor substrate but a potent inhibitor of rat and human neutrophil leukotriene A4 hydrolase. J Biol Chem. 1985 Sep 15;260(20):10966–10970. [PubMed] [Google Scholar]
  4. Ford-Hutchinson A. W. Leukotriene B4 in inflammation. Crit Rev Immunol. 1990;10(1):1–12. [PubMed] [Google Scholar]
  5. Griffin K. J., Gierse J., Krivi G., Fitzpatrick F. A. Opioid peptides are substrates for the bifunctional enzyme LTA4 hydrolase/aminopeptidase. Prostaglandins. 1992 Sep;44(3):251–257. doi: 10.1016/0090-6980(92)90018-o. [DOI] [PubMed] [Google Scholar]
  6. Haeggström J. Z., Wetterholm A., Medina J. F., Samuelsson B. Leukotriene A4 hydrolase: structural and functional properties of the active center. J Lipid Mediat. 1993 Mar-Apr;6(1-3):1–13. [PubMed] [Google Scholar]
  7. Haeggström J. Z., Wetterholm A., Vallee B. L., Samuelsson B. Leukotriene A4 hydrolase: an epoxide hydrolase with peptidase activity. Biochem Biophys Res Commun. 1990 Nov 30;173(1):431–437. doi: 10.1016/s0006-291x(05)81076-9. [DOI] [PubMed] [Google Scholar]
  8. Haeggström J., Rådmark O., Fitzpatrick F. A. Leukotriene A4-hydrolase activity in guinea pig and human liver. Biochim Biophys Acta. 1985 Jul 9;835(2):378–384. doi: 10.1016/0005-2760(85)90294-2. [DOI] [PubMed] [Google Scholar]
  9. Malfroy B., Kado-Fong H., Gros C., Giros B., Schwartz J. C., Hellmiss R. Molecular cloning and amino acid sequence of rat kidney aminopeptidase M: a member of a super family of zinc-metallohydrolases. Biochem Biophys Res Commun. 1989 May 30;161(1):236–241. doi: 10.1016/0006-291x(89)91586-6. [DOI] [PubMed] [Google Scholar]
  10. McGee J., Fitzpatrick F. Enzymatic hydration of leukotriene A4. Purification and characterization of a novel epoxide hydrolase from human erythrocytes. J Biol Chem. 1985 Oct 15;260(23):12832–12837. [PubMed] [Google Scholar]
  11. Minami M., Ohishi N., Mutoh H., Izumi T., Bito H., Wada H., Seyama Y., Toh H., Shimizu T. Leukotriene A4 hydrolase is a zinc-containing aminopeptidase. Biochem Biophys Res Commun. 1990 Dec 14;173(2):620–626. doi: 10.1016/s0006-291x(05)80080-4. [DOI] [PubMed] [Google Scholar]
  12. Mueller M. J., Samuelsson B., Haeggström J. Z. Chemical modification of leukotriene A4 hydrolase. Indications for essential tyrosyl and arginyl residues at the active site. Biochemistry. 1995 Mar 21;34(11):3536–3543. doi: 10.1021/bi00011a007. [DOI] [PubMed] [Google Scholar]
  13. Ohishi N., Izumi T., Minami M., Kitamura S., Seyama Y., Ohkawa S., Terao S., Yotsumoto H., Takaku F., Shimizu T. Leukotriene A4 hydrolase in the human lung. Inactivation of the enzyme with leukotriene A4 isomers. J Biol Chem. 1987 Jul 25;262(21):10200–10205. [PubMed] [Google Scholar]
  14. Orning L., Gierse J. K., Fitzpatrick F. A. The bifunctional enzyme leukotriene-A4 hydrolase is an arginine aminopeptidase of high efficiency and specificity. J Biol Chem. 1994 Apr 15;269(15):11269–11273. [PubMed] [Google Scholar]
  15. Orning L., Gierse J., Duffin K., Bild G., Krivi G., Fitzpatrick F. A. Mechanism-based inactivation of leukotriene A4 hydrolase/aminopeptidase by leukotriene A4. Mass spectrometric and kinetic characterization. J Biol Chem. 1992 Nov 15;267(32):22733–22739. [PubMed] [Google Scholar]
  16. Orning L., Jones D. A., Fitzpatrick F. A. Mechanism-based inactivation of leukotriene A4 hydrolase during leukotriene B4 formation by human erythrocytes. J Biol Chem. 1990 Sep 5;265(25):14911–14916. [PubMed] [Google Scholar]
  17. Orning L., Krivi G., Bild G., Gierse J., Aykent S., Fitzpatrick F. A. Inhibition of leukotriene A4 hydrolase/aminopeptidase by captopril. J Biol Chem. 1991 Sep 5;266(25):16507–16511. [PubMed] [Google Scholar]
  18. Orning L., Krivi G., Fitzpatrick F. A. Leukotriene A4 hydrolase. Inhibition by bestatin and intrinsic aminopeptidase activity establish its functional resemblance to metallohydrolase enzymes. J Biol Chem. 1991 Jan 25;266(3):1375–1378. [PubMed] [Google Scholar]
  19. Sun F. F., McGuire J. C. Metabolism of arachidonic acid by human neutrophils. Characterization of the enzymatic reactions that lead to the synthesis of leukotriene B4. Biochim Biophys Acta. 1984 Jun 6;794(1):56–64. doi: 10.1016/0005-2760(84)90297-2. [DOI] [PubMed] [Google Scholar]
  20. Vallee B. L., Auld D. S. Zinc coordination, function, and structure of zinc enzymes and other proteins. Biochemistry. 1990 Jun 19;29(24):5647–5659. doi: 10.1021/bi00476a001. [DOI] [PubMed] [Google Scholar]
  21. Wetterholm A., Haeggström J. Z. Leukotriene A4 hydrolase: an anion activated peptidase. Biochim Biophys Acta. 1992 Feb 12;1123(3):275–281. doi: 10.1016/0005-2760(92)90007-i. [DOI] [PubMed] [Google Scholar]
  22. Wetterholm A., Macchia L., Haeggström J. Z. Zinc and other divalent cations inhibit purified leukotriene A4 hydrolase and leukotriene B4 biosynthesis in human polymorphonuclear leukocytes. Arch Biochem Biophys. 1994 Jun;311(2):263–271. doi: 10.1006/abbi.1994.1236. [DOI] [PubMed] [Google Scholar]
  23. Wetterholm A., Medina J. F., Rådmark O., Shapiro R., Haeggström J. Z., Vallee B. L., Samuelsson B. Recombinant mouse leukotriene A4 hydrolase: a zinc metalloenzyme with dual enzymatic activities. Biochim Biophys Acta. 1991 Oct 25;1080(2):96–102. doi: 10.1016/0167-4838(91)90134-l. [DOI] [PubMed] [Google Scholar]

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