Leishmania amastigotes do not appear to alter the phagolysosomal pH (27), but rather possess membrane transport systems that are designed to exploit the acidic environment provided by the lysosome. Adaptation of amastigotes to high extracellular proton concentrations is strikingly evidenced by their metabolic behavior. Amastigotes optimally respire, metabolize glucose, and incorporate thymidine, uridine, and proline when bathed in medium between pH 4.0 and 5.5 (22). pH levels comparable to those within phagolysosomes. The effect of extracellular protons on the metabolic activity of amastigotes is probably due to increased nutrient transport through proton-coupled symporters, such as the glucose transporter (37,38). Proton influx is countered in both amastigotes and promastigotes by a plasmatemma proton pump, which is capable of generating a proton gradient across the plasma membrane (37; T. Glaser and A. Mukkada, personal communication). While the possible existence of other cation pumps has not been rigorously excluded, available evidence suggests that L. donotiani may rely solely on a proton pump to drive the uptake of nutrients and to regulate ion flow (37
An oligonucleotide probe was used to clone a cation-transporting ATPase gene from the genome of Leishmania donovani. The nucleotide sequence of the gene contained a 2,922-base-pair open reading frame that was predicted to encode a 107,406-dalton protein composed of 974 amino acids. The predicted L. donovani protein contained all the structural and functional domains expected to be present in a cation-transporting ATPase of the aspartyl phosphate class. The nucleotide sequence encoding the ATPase gene was duplicated in tandem in the parasite genome. Partial sequenation of the second member of the tandem repeat, which lay 2 kilobase pairs downstream of the ATPase gene, indicated that it was either identical to the first gene or very closely related to it. RNA homologous to either the ATPase gene or its adjacent relative was 5 kilobases in size and was approximately equally abundant in both promastigote and amastigote forms of the organism.
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