Identification and purification of a bovine liver mitochondrial NAD(+)-glycohydrolase.
Nonenzymatic ADP-ribosylation of mitochondrial proteins is thought to play a role in the regulation of Ca2+ efflux from mitochondria. It has been shown that intramitochondrial ADP-ribose is generated by a specific NAD(+)glycohydrolase, which catalizes hydrolysis of NAD+ to ADP-ribose and nicotinamide. We purified this enzyme from bovine liver mitochondrial membranes. The final preparation had a 1660-fold purified enzyme activity and contained a main protein band with an apparent molar mass of 32,000 in a SDS-polyacrylamide gel. The identity of this protein band with NAD(+)-glycohydrolase was verified by renaturation of its enzymatic activity. Partial amino acid sequence information was obtained from two enzyme fragments after proteolytic cleavage of the protein band in the SDS-polyacrylamide gel. Searches in protein databases revealed that an arginine ADP-ribosyl hydrolase harbours two stretches of amino acids that are highly similar to the partial NAD(+)-glycohydrolase sequences.
Institute of Biochemistry, University of Innsbruck, Austria.
J
J
Zhang
M
M
Ziegler
R
R
Schneider
H
H
Klocker
B
B
Auer
M
M
Schweiger
D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D002845:Chromatography; D004591:Electrophoresis, Polyacrylamide Gel; D008565:Membrane Proteins / Q000302:isolation & purification / Q000378:metabolism; D008930:Mitochondria, Liver / Q000201:enzymology*; D008969:Molecular Sequence Data; D009244:NAD+ Nucleosidase / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D017510:Protein Folding; D017421:Sequence Analysis; D013367:Submitochondrial Particles / Q000201:enzymology
D008565:Membrane Proteins; D009244:NAD+ Nucleosidase