Characterization and Localization of Acid Phosphatase Activity of Trypanosoma gambiense*

SYNOPSIS. Properties and cellular location of acid phosphatase in Trypanosoma gambiense were studied. Activity was found in both the sediment (32,000 ×g) and the supernatant of homogenates. Cenrifugation in 0.3 M sucrose showed activity principally in the lowspeed fraction (4,000 ×g). One min of sonication released most of this activity. Several phosphomonoesters were hydrolyzed at acid H's. Enzymatic activity was relatively specific for pyrophosphate and p‐nitrophenylphosphate at pH 3.6. At pH 5.2, purine and pyimidine nucleotide 5′‐triphosphates as well as adenosine di‐ and ono‐5′‐phosphates were hydrolyzed nonspecifically. Activity with yrophosphate at pH 3.6 had a temperature optimum of 60‐70 C while that for adenosine 5′‐triphosphate (pH 5.2) was 50 C. These ctivities of the sediment required no metal co‐factors and were inibited by Fe++, inhibition at the lower pH being greater. Glucose 6‐phosphate was hydrolyzed by the supernatant with maximum activity between pH 6.0 and 7.2 and a temperature optimum of 50 C. This pH range showed a broad plateau with 2 or 3 minor peaks. The hydrolysis of p‐nitrophenylphosphate showed a similar pH curve. In glucose 6‐phosphate hydrolysis, Mg++ was a required co‐factor but could be replaced by Ni++ or Co++. Ammonium sulfate fractionation precipitated most of the supernatant activity between 50 and 75% saturation. A modified Gomori technic produced spherical deposits of PbS thruout the cytoplasm of the intact cell. With the electron microscope, Pb phosphate deposition was observed in membrane‐bound vesicles (i.e., lysosomes) approximately 100‐150 mμ in diameter. These organelles were common in the region of the reservoir at the base of the flagellum. Acid phosphatase activity specific for glucose 6‐phosphate as substrate was localized within this basal pocket.