The Localization of Enzymes in Cells

Abstract

Summary 1. Enzymes and related substances may be localized in cells either, (a) by separation and analysis of cell parts, or (b) by application of light absorption methods (with or without staining) to living cells or tissue sections. 2. Separation of cell parts has been achieved by microdissection, by centrifuga‐tion (sea‐urchin eggs), by sectioning (neurones or centrifugally stratified Amoebae) and by bulk centrifugation of tissue suspensions. Highly sensitive micromethods, based on titration, dilatometry, or manometry, have been evolved for measuring the enzyme content of small fragments of protoplasm. 3. Precautions must be taken to minimize separation artefacts. In the bulk centrifugation techniques it is only possible to draw satisfactory conclusions when an enzyme remains associated with certain granules after repeated washing with physiological solutions. Even if an enzyme is constantly associated with, for instance, the mitochondria, it may occur only in certain of them, for they are not necessarily all identical. It is important (a) to distinguish between enzyme activity under the conditions existing in the cell and the maximum activity shown by an extract under optimal conditions, (b) to ensure that the total quantity of enzyme in the fractions separated equals that of the whole cell or tissue, (c) to distinguish between the concentration of, for instance, peptidase in any fraction and the proportion of the total cell peptidase present in the fraction. 4. The investigations which have most nearly satisfied these requirements have led to the following conclusions: (a) In various nuclei, dipeptidase, alkaline and acid phosphatase, arginase, uricase and esterase have been detected, all in lower absolute amounts than in the cytoplasm and all except alkaline phosphatase, arginase and uricase in lower concentration. Chromosome‐like threads can be centrifuged from sperm and suspensions of somatic tissues; after removal of desoxyribonucleohistone, there remains a residual chromosome containing ribonucleic acid, alkaline phosphatase and a tryptophane‐containing protein as well as other components, (b) The major part of the cytochrome oxidase, of the succinic dehydrogenase and of several enzymes concerned in fatty acid oxidation and in the Krebs tricarboxylic acid cycle is present in the mitochondria in rat‐liver cells. Mitochondria contain neither dipeptidase nor catalase in sea‐urchin eggs and Amoebae, but in the latter they appear to be associated with amylase activity, (c) Microsomes (cytoplasmic particles 50–200 m/j. in diameter) have been found to contain dipeptidase, catalase, phosphatases, ribonuclease, about half the esterase and half the ribonucleic acid in adult liver cells, (d) Much of the catalase, carboxylase and lactic acid dehydrogenase in yeast, of the glycolytic activity in rat liver and of the ribonucleic acid in embryonic tissues and tumours remains in the supernatant after centrifugation for 1–2 hr. at 18,000 g., whether because these substances are freely dissolved in the protoplasm, or bec...

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