Research Project
3454419
Polyamine synthesis is eucaryotic cells traditionally involves the generation of small levels of amines for use as cofactors in cell division and synthesis of macromolecules. However, there are strong indications that in the urogenital parasite Trichomonas vaginalis these amines may be important in: a) energy metabolism; b) conditioning the environment; c) as oxidative substrate. THe proposed work is based on preliminary data indicating a major difference in synthesis and metabolism of polyamines between parasite and host. Ornithine, the immediate precursor of putrescine is synthesized via the arginine dihydrolase pathway generating ATP. Internal putrescine levels rise significantly to 13mM in organisms grown in semi-defined medium resulting in its eventual excretion into the medium. While the putrescine analog, cadaverine, is also synthesized, the major polyamine spermidine and spermine are present only in low levels. Polyamine synthetic enzyme, S-adenosylmethionine decarboxylase, spermidine and spermine synthetases appear to be lacking. A spermidine specific, aminoguanidine-sensitive polyamine oxidase is present in a metronidazole-sensitive but not in a refractory strain. These findings point to unusual polyamine metabolism in this organism - one that yields ATP production from amino acids in a low carbohydrate environment and that produces excessive putrescine. These aspects of parasite metabolism differ significantly from the host and present potential targets for chemotherapy. Polyamine metabolism in T. vaginalis will be investigated in metronidazole sensitive and refractory strains under defined growth conditions to determine: (1) Whether under natural substrate conditions in which the carbohydrate supply is limiting, the arginine dihydrolase pathway leading to production of ornithine and ATP via carbamoyl phosphate, is critical for energy metabolism aerobically and anaerobically. (2) The types of polyamines, their routes of synthesis and precursors and the effect on growth of amino acid analogs (e.g. difluoromethylornithine) which inhibits polyamine biosynthesis. (3) Whether sensitivity to metronidazole is related to the presence of an amino-guanidine-sensitive, strain-specific polyamine oxidase activity. (4) Does extensive putrescine production and excretion serve only to allow high metabolic flow through the arginine dihydrolase pathway or does the excess putrescine serve as growth factor and/or have some other role in conditioning of the environment.
