Identification and Characterization of Two Isoenzymes of Methionine γ-Lyase from Entamoeba histolytica

A key enzyme of sulfur-amino acid degradation in an anaerobic parasitic protist that lacks forward and reverse trans-sulfuration pathways

Masaharu Tokoro, Takashi Asai, Seiki Kobayashi, Tsutomu Takeuchi, Tomoyoshi Nozaki

Research output: Contribution to journalArticle

90 Citations (Scopus)

Abstract

To better understand the metabolism of sulfur-containing amino acids, which likely plays a key role in a variety of cell functions, in Entamoeba histolytica, we searched the genome data base for genes encoding putative orthologs of enzymes known to be involved in the metabolism. The search revealed that E. histolytica possesses only incomplete cysteine-methionine conversion pathways in both directions. Instead, this parasite possesses genes encoding two isoenzymes of methionine γ-lyase (EC 4.4.1.11, EhMGL1/2), which has been implicated in the degradation of sulfur-containing amino acids. The two amebic MGL isoenzymes, showing 69% identity to each other, encode 389- and 392-amino acid polypeptides with predicted molecular masses of 42.3 and 42. 7 kDa and pIs of 6.01 and 6.63, respectively. Amino acid comparison and phylogenetic analysis suggested that these amebic MGLs are likely to have been horizontally transferred from the Archaea, whereas an MGL from another anaerobic protist Trichomonas vaginalis has MGL isotypes that share a common ancestor with bacteria. Enzymological and immunoblot analyses of the partially purified native amebic MGL confirmed that both of the MGL isotypes are expressed in a comparable amount predominantly in the cytosol and form a homotetramer. Recombinant EhMGL1 and 2 proteins catalyzed degradation of L-methionine, DL-homocysteine, L-cysteine, and O-acetyl-L-serine to form α-keto acid, ammonia, and hydrogen sulfide or methanethiol, whereas activity toward cystathionine was negligible. These two isoenzymes showed notable differences in substrate specificity and pH optimum. In addition, we showed that EhMGL is an ideal target for the development of new chemotherapeutic agents against amebiasis by demonstrating an amebicidal effect of the methionine analog trifluoromethionine on trophozoites in culture (IC 50 18 μM) and that this effect of trifluoromethionine was completely abolished by the addition of the MGL-specific inhibitor DL-propargylglycine.

Original languageEnglish
Pages (from-to)42717-42727
Number of pages11
JournalJournal of Biological Chemistry
Volume278
Issue number43
DOIs
Publication statusPublished - 2003 Oct 24

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Sulfur Amino Acids
Entamoeba histolytica
Lyases
Methionine
Isoenzymes
Amino Acids
Degradation
Gene encoding
Enzymes
Sulfur
Metabolism
Cysteine
Cystathionine
Keto Acids
Trichomonas vaginalis
Amebiasis
Trophozoites
Hydrogen Sulfide
Archaea
Molecular mass

ASJC Scopus subject areas

  • Biochemistry

Cite this

Identification and Characterization of Two Isoenzymes of Methionine γ-Lyase from Entamoeba histolytica : A key enzyme of sulfur-amino acid degradation in an anaerobic parasitic protist that lacks forward and reverse trans-sulfuration pathways. / Tokoro, Masaharu; Asai, Takashi; Kobayashi, Seiki; Takeuchi, Tsutomu; Nozaki, Tomoyoshi.

In: Journal of Biological Chemistry, Vol. 278, No. 43, 24.10.2003, p. 42717-42727.

Research output: Contribution to journalArticle

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