Pyruvate decarboxylase, the target for omeprazole in metronidazole-resistant and iron-restricted Tritrichomonas foetus

Abstract

The substituted benzimidazole omeprazole, used for the treatment of human peptic ulcer disease, inhibits the growth of the metronidazole-resistant bovine pathogen Tritrichomonas foetus in vitro (MIC at which the growth of parasite cultures is inhibited by 50%, 22 microg/ml [63 microM]). The antitrichomonad activity appears to be due to the inhibition of pyruvate decarboxylase (PDC), which is the key enzyme responsible for ethanol production and which is strongly upregulated in metronidazole-resistant trichomonads. PDC was purified to homogeneity from the cytosol of metronidazole-resistant strain. The tetrameric enzyme of 60-kDa subunits is inhibited by omeprazole (50% inhibitory concentration, 16 microg/ml). Metronidazole-susceptible T. foetus, which expresses very little PDC, is only slightly affected. Omeprazole has the same inhibitory effect on T. foetus cells grown under iron-limited conditions. Similarly to metronidazole-resistant cells, T. foetus cells grown under iron-limited conditions have nonfunctional hydrogenosomal metabolism and rely on cytosolic PDC-mediated ethanol fermentation.

FIG. 1.

Structures of thiamine (a), omeprazole (b), and metronidazole (c).

FIG. 2.

(A) SDS-PAGE of PDC fractions during purification steps from metronidazole-resistant T. foetus cytosol. Lanes: 1, molecular weight standards; 2, cytosol; 3, fractions after butyl HIC column chromatography; 4, fractions obtained after hydroxyapatite column chromatography; 5, fractions obtained after UNO Q 1 column chromatography; 6, fractions obtained after Bio-Prep 1000 column chromatography. Proteins were stained with Coomassie brilliant blue. (B) Immunodetection of PDC in the cytosolic fractions of T. foetus. Lanes: 1, metronidazole-resistant strain; 2, metronidazole-sensitive strain; 3, metronidazole-sensitive strain grown under iron-limited conditions.

FIG. 3.

Effect of omeprazole on activity of purified PDC from metronidazole-resistant T. foetus. The data represent the means ± standard deviations of five measurements.