1. Studies were carried out with pure lactate dehydrogenase isoenzymes C4 (LDH isoenzyme X), B4, (LDH isoenzyme 1) and A4 (LDH isoenzyme 5) isolated from mouse testis, heart and muscle tissue respectively; with LDH isoenzyme X purified from pigeon testes and with crude lysates of spermatozoa from man, bull and rabbit. 2. LDH isoenzyme X from all species showed greater ability than the other isoenzymes to catalyse the NAD+-linked interconversions of 2-oxobutanoate into 2-hydroxybutanoate and of 2-oxopentanoate into 2-hydroxypentanoate. 3. Mouse LDH isoenzyme X presented the broadest spectrum of substrate specificity. It exhibited very similar Km values for a variety of 2-oxo acids: 2-oxopropanoate (pyruvate), 2-oxobutanoate, 2-oxo-3-methylbutanoate, 2-oxopentanoate, 2-oxo-3-methylpentanoate, 2-oxo-4-methylpentanoate, 2-oxohexanoate and 2-oxo-3-phenylpropanoate (phenylpyruvate). The corresponding 2-hydroxy acids were also readily utilized in the reverse reaction. A strong inhibition by substrate and product was demonstrated for the direct reaction. 4. Intracellular distribution of LDH isoenzyme X was investigated in mouse testes. LDH isoenzyme X activity was located in the fraction of “heavy mitochondria” and in the soluble phase. 5. A possible functional role for LDH isoenzyme X is proposed: the redox couple-2-oxo acid-2-hydroxy acid could integrate a shuttle system transferring reducing equivalents from cytoplasm to mitochondria.
Gossypol, a phenolic compound isolated from the cotton plant, is a powerful inhibitor of nicotinamide adenine dinucleotide-linked enzymes (alpha-hydroxyacid dehydrogenase and malate dehydrogenase) of Trypanosoma cruzi, the parasite that causes Chagas' disease. Parasites at the epimastigote stage that were incubated for 5 minutes with 100 micromolar gossypol were completely immobilized. Concentrations of gossypol as low as 0.01 micromolar markedly reduced the growth rate of T. cruzi in culture.
Isolates of Trypanosoma cruzi from human patients, domestic and sylvatic animals and vector insects were obtained in different areas of Argentina. Electrophoretic patterns of enzymes from extracts of 95 isolates were analysed. On the basis of zymograms providing information on 10 loci, 12 zymodemes are described according to their genotypes. Data presented show fixed heterozygosity, absence of segregation of genotypes, significant departures from Hardy-Weinberg equilibrium, and over-represented genotypes. This evidence supports the hypothesis that sexual reproduction is very restricted or absent in this parasite. The proportion of polymorphic loci is 80%. The expected mean heterozygosity per locus (He) is 0.43, while the observed value (Ho) is 0.24. Differences between these values may be explained by accepting a basically clonal structure for T. cruzi. The data matrix of 12 zymodemes using 28 characters was analysed using a Wagner parsimony algorithm. Two equally most parsimonious unrooted trees were generated; both have 39 steps. The results show clusters clearly separated according to the geographical origin of the stocks. There are some indications of some correlations between genetic composition of the parasite and the clinical picture of the infection in human patients.
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