Malaria parasites, unable to synthesize purine de novo, use host-derived hypoxanthine preferentially as purine source. In a previous study (1990. J. Biol. Chem. 265:6562-6568), we noted that xanthine oxidase rapidly and completely depleted hypoxanthine in human erythrocytes, not by crossing the erythrocyte membrane, but rather by creating a concentration gradient which facilitated hypoxanthine efilux. We therefore investigated the ability of xanthine oxidase to inhibit growth of FCR-3, a chloroquine-resistant strain of Plasmodium falciparum in human erythrocytes in vitro. Parasites were cultured in human group O erythrocytes in medium supplemented, as required, with xanthine oxidase or chloroquine. Parasite viability was assessed by uptake of radiolabeled glycine and adenosine triphosphate-derived purine into protein and nucleic acid, respectively, by nucleic acid accumulation, by L-lactate production, and by microscopic appearance. On average, a 90% inhibition of growth was observed after 72 h of incubation in 20 mU/ ml xanthine oxidase. Inhibition was notably greater than that exerted by i0' M chloroquine (< 10%) over a comparable period. The IC5o for xanthine oxidase was estimated at 0.2 mU/ ml, compared to 1.5 X i0' M for chloroquine. Inhibition was completely reversed by excess hypoxanthine, but was unaffected by oxygen radical scavengers, including superoxide dismutase and catalase. The data confirms that a supply of hostderived hypoxanthine is critical for nucleic acid synthesis in P. falciparum, and that depletion of erythrocyte hypoxanthine pools with xanthine oxidase offers a novel approach to treatment of chloroquine-resistant malaria infection in humans. (J.
The effect of pH, P02, and inorganic phosphate on the uptake and metabolism of hypoxanthine by erythrocytes has been studied. Uptake of hypoxanthine and accumulation of inosine 5'-monophosphate (IMP) were markedly increased at acid pH, high external phosphate concentrations, and low Po2. Release of accumulated IMP as hypoxanthine occurred at alkaline pH values and low external phosphate concentrations. Conditions favoring IMP accumulation gave rise, in the absence of hypoxanthine, to a corresponding increase in 5'-phosphoribosyl-1-pyrophosphate. Intracellular phosphate concentrations were markedly pH dependent and a model is presented whereby hypoxanthine uptake and release are controlled by intracellular concentrations of inorganic phosphate and 2,3-bisphosphoglycerate. These allosteric effectors influence, in opposing ways, two enzymes governing IMP accumulation, namely 5'-phosphoribosyl-l-pyrophosphate synthetase and 5'-nucleotidase. These metabolic properties suggest that the erythrocyte could play a role in the removal of hypoxanthine from anoxic tissue.
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