Distribution of Acid Phosphatase in Liver

Of several methods that are potentially available for determining the intracellular distribution of acid phosphatase and other enzymes, the chief ones currently in use are ultracentrifugal separation and histochem-ical examination. Each of these has its disadvantages and advantages, some of which have already been indicated. At this point we will consider the quantitative ultracentrifugal methods. 

As may be seen, the light mitochondrial fraction, as obtained by de Duve et al. (DIO), contained only 40.7% of the acid phosphatase of the rat liver. Starting with this fraction, Sawant et al. (S4) attempted 

The yield of acid phosphatase was 11%, as compared with 30% for aryl sulfatase and 15% for ribonuclease. Obviously, in the attempt to obtain pure preparations of intracellular components, it is inevitable that losses be encountered. Such procedures are therefore not suited for obtaining an estimate of the quantitative distribution of intracellular components. Approximately 5-9% of the lysosomal enzymes—aryl sulfatase, acid phosphatase, and ribonuclease—were present in the free form. The remaining 91-95% of the activities of these enzymes were in the latent form and required alteration of the permeability or disruption of the lysosomal membrane to become active. 

It has been estimated that approximately 70-80% of the acid phosphatase in rat liver can be recovered in the lysosomal fraction, the remainder being distributed between the soluble fraction and other sub-cellular fractions (D9, S15). The question therefore arises concerning the extent to which this remainder is derived from lysosomes broken during the fractionation procedure or whether some acid phosphatase is actually localized in subcellular structures, other than lysosomes. 

The lysosomal and the soluble fractions of acid phosphatase were compared in several other ways. The values (milf) for the lysosomal fractions on various substrates were -glycerophosphate, 1.6 fructose 1,6-diphosphate, 2.0 p-nitrophenyl phosphate, 1.6 AMP, 0.43 they were not significantly different from those obtained with the soluble fraction. The pH-activity curves with these substrates were similar for the two fractions. Inhibition of phosphatase activity of the lysosomal and soluble fractions occurred at approximately the same concentration of fluoride or n- (+) -tartrate when j8-glycerophosphate, AMP, or fructose 1,6-diphosphate were used as substrates. However, with p-nitrophenyl phosphate 

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