The characteristics and intracellular localization of Ehrlich ascites tumor cell glutaminase were investigated. It was found that these cells possess only the phosphate-dependent glutaminase isoenzyme, which requires 50 mm inorganic phosphate for maximum activation. The Km value of the enzyme for glutamine was 4.5 mm, and the optimum pH was between 8.0 and 8.5. Differential centrifugation of the cell homogenate revealed that glutaminase strictly follows the distribution of glutamate dehydrogenase, indicating that this enzyme has an exclusively mitochondrial localization. Disintegration of the mitochondria led to the complete inactivation of glutaminase, but the addition of borate protected the enzyme when detergents were used. The treatment of the mitochondria with digitonin followed by centrifugation showed that glutaminase is located in the mitochondrial matrix.
This work was supported by The Community for Scientific Research of Autonomous Province of Vozvodina, Grant 01-385/1.