Subcellular fractionation by differential and zonal centrifugation of aerobically grown glucose-de-repressed Saccharomyces carlsbergensis.
1. Homogenates were prepared from sphaeroplasts of aerobically grown glucose-de-repressed Saccharomyces carlsbergensis and the distributions of marker enzymes were investigated after differential centrifugation. Cytochrome c oxidase and cytochrome c were sedimented almost completely at 10(5)g-min, and this fraction also contained 37% of the catalase, 27% of the acid p-nitrophenyl phosphatase, 53 and 54% respectively of the NADH- and NADPH-cytochrome c oxidoreductases. 2. Zonal centrifugation indicated complex density distributions of the sedimentable portions of these enzymes and of adenosine triphosphatases and suggested the presence of two mitochondrial populations, as well as a bimodal distribution of peroxisomes and heterogeneity of the acid p-nitrophenyl phosphatase-containing particles. 3. Several different adenosine triphosphatases were distinguished in a post-mitochondrial supernatant that contained no mitochondrial fragments; these enzymes varied in their sensitivities to oligomycin and ouabain and their distributions were different from those of pyrophosphatase, adenosine phosphatase and adenosine pyrophosphatase. 4. The distribution of NADPH-cytochrome c oxidoreductase demonstrated that it cannot be used in S. carlsbergensis as a specific marker enzyme for the microsomal fraction. Glucose 6-phosphatase, inosine pyrophosphatase, cytochrome P-450 and five other enzymes frequently assigned to microsomal fractions of mammalian origin were not detected in yeast under these growth conditions.
D000251:Adenosine Triphosphatases / Q000032:analysis; D000332:Aerobiosis; D002374:Catalase / Q000032:analysis; D002458:Cell Fractionation; D002499:Centrifugation, Density Gradient; D002501:Centrifugation, Zonal; D003580:Cytochromes / Q000032:analysis; D003576:Electron Transport Complex IV / Q000032:analysis; D000431:Ethanol / Q000378:metabolism; D005947:Glucose / Q000378:metabolism; D005952:Glucose-6-Phosphatase / Q000032:analysis; D007288:Inosine; D008854:Microscopy, Electron; D009243:NAD; D009249:NADP; D010088:Oxidoreductases / Q000032:analysis; D010744:Phosphoric Monoester Hydrolases / Q000032:analysis; D011523:Protoplasts; D011755:Pyrophosphatases / Q000032:analysis; D012440:Saccharomyces / Q000166:cytology* / Q000201:enzymology / Q000254:growth & development / Q000378:metabolism; D013347:Subcellular Fractions / Q000201:enzymology
D003580:Cytochromes; D009243:NAD; D000431:Ethanol; D009249:NADP; D007288:Inosine; D010088:Oxidoreductases; D002374:Catalase; D003576:Electron Transport Complex IV; D010744:Phosphoric Monoester Hydrolases; D005952:Glucose-6-Phosphatase; D000251:Adenosine Triphosphatases; D011755:Pyrophosphatases; D005947:Glucose