Tissue fractionation studies. 6. Intracellular distribution patterns of enzymes in rat-liver tissue

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Abstract
The distribution of a number of enzymes was investigated according to a new fractionation scheme, in which the classical mitochondria are divided into 2 subtractions. The distribution patterns were compared to those of cytochrome oxidase, acid phosphatase and glucose-6-phos-phatase, which served as reference enzymes. The data were interpreted in the light of these comparisons and by additional experiments. The distribution patterns of succinate cytochrome c reductase and of rhodanese followed that of cytochrome oxidase; it was concluded that these systems belong to the mitochondria. The DPNH- and TPNH-cytochrome c reductase activities showed complex distributions, reflecting the existence of 2 distinct pairs of systems, associated with the mitochondria and the microsomes and differing in their susceptibility to activation by distilled water and inhibition by antimycin A. About 40% of the total fumarase activity was found in the mitochondria, the remainder recovered partly in the final supernatant, partly in the final supernatant, partly in the microsomes, possibly in the latter case as the result of an adsorption artifact. Ribonuclease, deoxyribonuclease, cathepsin and the major part of [beta]-glucuronidase showed distributions analogous to that of acid phosphatase. They also resembled this enzyme in being practically unreactive towards their respective substrates in intact preparations and were released in a parallel fashion from granules subjected to graded activation by a variety of means. Results indicated that 4 hydrolases belong to the same distinct group of granules previously shown to contain acid phosphatase. The name lysosomes was proposed for these granules. The microsomes contained a 20% excess of B-glucuronidase, which differed from the remaining activity by a less acid pH optimum. A 2nd enzyme species appears to be involved, but may possibily be an interaction between adsorbed enzyme and ribo-nucleic acids. Uricase had a unique distribution, indicating that it is attached either to the insoluble framework of lysosomes or to a 4th distinct group of granules with the properties of large microsomes. Many of these results support assumptions that distinct enzymic species have single intracellular locations and that granules of a given class are enzymically homogeneous. Some conflicting evidence has also been brought to light.