Characterization of hydrosoluble and detergent-solubilized forms of mitochondrial NAD+ glycohydrolase from bovine liver.
Treatment of isolated bovine liver mitochondria with either detergents or a crude pancreatic lipase, steapsin, resulted in solubilization of NAD+ glycohydrolase (NADase) activity. The two forms of this enzyme can be visualized directly in SDS-polyacrylamide gels (PAGs) by a fluorescence assay utilizing 1,N6-etheno-NAD+ (epsilon-NAD+) as substrate. Only a slight difference of about 2,000 in the apparent molecular masses was detected. Values of 28,000 and 30,000 for the steapsin- and detergent-solubilized enzyme, respectively, were estimated. The catalytic properties as well as the dependence on temperature, pH, and ionic strength were found to be similar for both forms of the enzyme. One important difference regarding their sensitivity against bivalent metal ions was observed. While the detergent-solubilized NADase was activated in the presence of, for example, Zn++, and inhibited by EDTA, the truncated enzyme seemed to be unaffected under these conditions.
Jorcke
D
D
Freie Universität Berlin, Institut f. Biochemie, Germany.
Ziegler
M
M
Schweiger
M
M
D000818:Animals; D002417:Cattle; D002855:Chromatography, Thin Layer; D003902:Detergents / Q000494:pharmacology; D004229:Dithiothreitol / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D004791:Enzyme Inhibitors / Q000494:pharmacology; D008049:Lipase / Q000494:pharmacology; D008274:Magnesium / Q000494:pharmacology; D008565:Membrane Proteins / Q000737:chemistry / Q000187:drug effects / Q000302:isolation & purification / Q000378:metabolism; D008623:Mercaptoethanol / Q000494:pharmacology; D008930:Mitochondria, Liver / Q000201:enzymology*; D009243:NAD / Q000031:analogs & derivatives / Q000378:metabolism; D009244:NAD+ Nucleosidase / Q000737:chemistry / Q000187:drug effects / Q000302:isolation & purification / Q000378:metabolism*; D010179:Pancreas / Q000201:enzymology; D012995:Solubility; D014867:Water; D015032:Zinc / Q000494:pharmacology
D003902:Detergents; D004791:Enzyme Inhibitors; D008565:Membrane Proteins; D014867:Water; D009243:NAD; C011038:nicotinamide 1,N(6)-ethenoadenine dinucleotide; D008623:Mercaptoethanol; D008049:Lipase; D009244:NAD+ Nucleosidase; D008274:Magnesium; D015032:Zinc; D004229:Dithiothreitol