CHARACTERIZATION OF 2 FORMS OF POLY(ADP-RIBOSE) GLYCOHYDROLASE IN GUINEA-PIG LIVER

A poly(ADP-ribose) glycohydrolase from guinea pig liver cytoplasm has been purified approximately 45 000-fold to apparent homogeneity. The cytoplasmic poly(ADP-ribose) glycohydrolase designated form II differed in several respects from the nuclear poly(ADP-ribose) glycohydrolase I (M(r), = 75 500) previously purified from the same tissue (Tanuma et al., 1986a). The purified glycohydrolase II consists of a single polypeptide with M(r) of 59 500 estimated by a sodium dodecyl sulfate-polyacrylamide gel. A native M(r) of 57 000 was determined by gel permeation. Peptide analysis of partial proteolytic degradation of glycohydrolases II and I with Staphylococcus aureus V8 protease revealed that the two enzymes were structurally different. Amino acid analysis showed that glycohydrolase II had a relatively low proportion of basic amino acid residues as compared with glycohydrolase I. Glycohydrolase II and I were acidic proteins with isoelectric points of 6.2 and 6.6, respectively. The optimum pH for glycohydrolases II and I were around 7.4 and 7.0, respectively. The K(m) value for (ADP-ribose)n (average chain length n = 15) and the V(max) for glycohydrolase II were 4.8-mu-M and 18-mu-mol of ADP-ribose released from (ADP-ribose)n.min-1.(mg of protein)-1, respectively. The K(m) was about 2.5 times higher, and V(max), 2 times lower, than those observed with glycohydrolase 1. Unlike glycohydrolase I, glycohydrolase II was inhibited by monovalent salts. ADP-ribose and cAMP inhibited glycohydrolase II more strongly than glycohydrolase I. These results suggest that eukaryotic cells contain two distinct forms of poly(ADP-ribose) glycohydrolase exhibiting differences in properties and subcellular localization.