Alternative 2-keto acid oxidoreductase activities in Trichomonas vaginalis

• Published in: Molecular and biochemical parasitology 1999-01, Vol.98 (2), p.203-214
• Main Authors: Brown, David M., Upcroft, Jacqueline A., Dodd, Helen N., Chen, Nanhua, Upcroft, Peter
• Format: Article
• Language: English
• Subjects: 2-keto acid oxidoreductase; Alternative energy production; Animals; Antitrichomonal Agents - pharmacology; Cell Compartmentation; Drug Resistance; Energy Metabolism; Isoenzymes; Ketone Oxidoreductases - biosynthesis; Ketone Oxidoreductases - genetics; Ketone Oxidoreductases - isolation & purification; Localization; Metronidazole - pharmacology; Metronidazole resistance; Pyruvate Synthase; Pyruvate:ferredoxin oxidoreductase; Pyruvate:Indolepyruvate; RNA, Messenger - isolation & purification; RNA, Protozoan - isolation & purification; Solubility; Subcellular Fractions - enzymology; Trichomonas vaginalis; Trichomonas vaginalis - enzymology; α-ketoglutarate
• ISSN: 0166-6851; 1872-9428
• DOI: 10.1016/S0166-6851(98)00169-8

We have induced high levels of resistance to metronidazole (1 mM or 170 μg ml −1) in two different strains of Trichomonas vaginalis (BRIS/92/STDL/F1623 and BRIS/92/STDL/B7708) and have used one strain to identify two alternative T. vaginalis 2-keto acid oxidoreductases (KOR) both of which are distinct from the already characterised pyruvate:ferredoxin oxidoreductase (PFOR). Unlike the characterised PFOR which is severely down-regulated in metronidazole-resistant parasites, both of the alternative KORs are fully active in metronidazole-resistant T. vaginalis. The first, KOR1, localized in all membrane fractions but predominantly in the hydrogenosome fraction, is soluble in Triton X-100 and the second, KOR2, is extractable in 1 M acetate from membrane fractions of metronidazole-resistant parasites. PFOR and both KOR1 and KOR2 use a broad range of 2-keto acids as substrates (pyruvate, α-ketobutyrate, α-ketomalonate), including the deaminated forms of aromatic amino acids (indolepyruvate and phenylpyruvate). However, unlike PFOR neither KOR1 or KOR2 was able to use α-ketoglutarate. Deaminated forms of branched chain amino acids (α-ketoisovalerate) were not substrates for T. vaginalis KORs. Since KOR1 and KOR2 do not apparently donate electrons to ferredoxin, and are not down-regulated in metronidazole-resistant parasites, we propose that KOR1 and KOR2 provide metronidazole-resistant parasites with an alternative energy production pathway(s) which circumvents metronidazole activation.