[129] Polymerization of the adenosine 5′-diphosphate-ribose moiety of NAD

Publisher Summary This chapter discusses the polymerization of the adenosine 5'-diphosphate-ribose moiety of nicotinamide adenine dinucleotide (NAD). The polymerization is catalyzed by certain aggregated enzyme preparations. This enzymatic polymerization is accompanied by a release of the nicotinamide moiety of NAD. No energy source is required for the polymerization as nicotinamide inhibits the reaction. Under optimal conditions, about 30% of NAD added as a substrate is converted to poly (ADP-Rib) adenosine diphosphate ribose. Poly (ADP-Rib) can be purified by the following steps: acetate buffer-ethanol precipitation; pronase digestion; phenol extraction; pancreatic RNase and DNase digestion; and gel filtration on Sephadex. For further purification, digestion with micrococcal nuclease and spleen phosphodiesterase is applied. Enzymatic formation of ADP-ribose polymer involves enzyme preparation. To follow the enzymatic formation poly (ADP-Rib), the use of NAD labeled with radioactive isotope is recommended. Ado (P)-Rib-P is purified from the phosphodiesterase hydrolyzate of polymer on Dowex l-X8 (200–400 mesh) formate column chromatography. The properties of poly (ADP-Rib) are also discussed.