This chapter discusses the preparation and properties of α- (NAD+) nicotinate adenine dinucleotide and α-NADH. α-NAD+ is separated from a commercial NAD+ preparation by employing an ion-exchange chromatographic method. α-NADH is prepared by chemical reduction of α-NAD+. α-NADH is prepared by chemical reduction of α-NAD+ with hydro-sulfite and isolated as the barium salt. The existence of several enzyme systems in mammals catalyzing the transfer of electrons from α-NADH to electron acceptors such as cytochrome c, 2, 6-dichlorophenolindophenol, O2 is described. Protein concentrations are estimated by using crystalline bovine plasma albumin as a standard. The α-NADH-cytochrome c reductase activity of the original homogenates is detected in the microsomal fraction, whereas the activity of the mitochondrial fraction is only 12%. On the other hand, most of the α-NADH-2, 6-DCPIP reductase activity is detected in the soluble fraction. Diaphorase activities with α- and β-NADH at each purification step is summarized. Purification of diaphorase (α-NADH-2,6-DCPIP reductase) are carried out from fresh homogenized pig heart.
|Number of pages||5|
|Journal||Methods in enzymology|
|Publication status||Published - 1971 Jan 1|
ASJC Scopus subject areas
- Molecular Biology