Analytical fractionation of homogenates from cultured rat embryo fibroblasts.
Homogenates of cultured rat embryo fibroblasts have been assayed for acid phosphatase, N-acetyl-beta-glucosaminidase, cathepsin D, acid deoxyribonuclease, cytochrome oxidase, NADH cytochrome c reductase, 5'-nucleotidase, inosine diphosphatase, acid pyrophosphatase, neutral pyrophosphatase, esterase, catalase, cholesterol, and RNA. The validity of the assay conditions was checked. Neutral pyrophosphatase is a readily soluble enzyme. Acid hydrolases, except acid pyrophosphatase, are particle-bound enzymes, which exhibit a high degree of structural latency. They are activated and solubilized in a parallel fashion by mechanical treatments and tensio-active agents. Catalase is also particle-bound and latent; activating conditions stronger than those for hydrolases are required to activate the enzyme. Acid pyrophosphatase, 5'-nucleotidase and inosine diphosphatase are firmly particle-bound, but not latent; they are not easily solubilized. In differential and isopycnic centrifugation, the latent hydrolases, cytochrome oxidase and catalase dissociate largely from each other; this suggests the occurrence of lysosomes and peroxisome-like structures besides mitochondria. The distribution patterns of 5'-nucleotidase and cholesterol are largely similar; digitonin influences their equilibrium density to the same extent; these two constituents are thought to be related to the plasma membrane. Inosine diphosphatase and acid pyrophosphatase are also partially associated with the plasma membrane, although some part of these enzymic activities probably belongs to other structures. NADH cytochrome c reductase is associated partly with the endoplasmic reticulum, partly with mitochondria.
D000135:Acid Phosphatase / Q000032:analysis; D000818:Animals; D002374:Catalase / Q000032:analysis; D002403:Cathepsins / Q000032:analysis; D002462:Cell Membrane / Q000201:enzymology; D002499:Centrifugation, Density Gradient; D003579:Cytochrome Reductases / Q000032:analysis; D003594:Cytoplasmic Granules / Q000201:enzymology; D003851:Deoxyribonucleases / Q000032:analysis; D003576:Electron Transport Complex IV / Q000032:analysis; D004622:Embryo, Mammalian; D004721:Endoplasmic Reticulum / Q000201:enzymology; D004950:Esterases / Q000032:analysis; D005260:Female; D005347:Fibroblasts / Q000201:enzymology* / Q000648:ultrastructure; D006596:Hexosaminidases / Q000032:analysis; D006651:Histocytochemistry; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008247:Lysosomes / Q000201:enzymology; D008854:Microscopy, Electron; D008928:Mitochondria / Q000201:enzymology; D009708:Nucleotidases / Q000032:analysis; D011247:Pregnancy; D011755:Pyrophosphatases / Q000032:analysis; D051381:Rats
D002374:Catalase; D003579:Cytochrome Reductases; D003576:Electron Transport Complex IV; D003851:Deoxyribonucleases; D004950:Esterases; D009708:Nucleotidases; D000135:Acid Phosphatase; D006596:Hexosaminidases; D002403:Cathepsins; D011755:Pyrophosphatases