Diphosphopyridine nucleotide pyrophosphatase.

Enzymatic splitting of diphosphopyridine nucleotide (DPN)’ by microorganisms, plants, and animal tissues has been known for many years, but the precise nature of this reaction is not well understood (1). Ohlmeyer (2) first observed that adenylic acid (5-phosphoadenosine) appeared when DPN was incubated with yeast maceration juice. Later Heiwinkel (3) identified adenylic acid as a product of the action of sweet almond press juice on DPN. Although free nicotinamide did not appear, the presumed nicotinamide mononucleotide could not be isolated. Das and von Euler (4, 5) found inorganic orthophosphate as a product of DPN destruction in animal tissues, but there was no indication as to the mechanism of this reaction. Handler and Klein (6) recovered free nicotinamide quantitatively as a product of DPN destruction by brain, liver, kidney, and muscle preparations from rabbits, rats, and dogs, and concluded that the nicotinamide-ribose linkage was the primary and principal site of cleavage in these tissues. In the course of studies of oxidative phosphorylation, we observed that washed particles of rabbit kidney converted added DPN to adenosine triphosphate (ATP). In the absence of an oxidizable substrate to provide an energy source for phosphorylation, adenylic acid was formed in place of ATP. The other product of DPN splitting was tentatively identified, after partial purification, as nicotinamide ribose phosphate. These and related observat.ions make it probable that in such rabbit kidney preparations cleavage of the pyrophosphate bond is the predominant mechanism of DPN splitting. Almost all of the DPN-splitting activity of rabbit brain is due to nucleosidase activity.