Developmental and biochemical characteristics of the cardiac membrane-bound arginine-specific mono-ADP-ribosyltransferase.

ADP-ribosylation of protein in heart membrane preparations has been shown to be present in adult tissue but absent from early neonate tissue [Piron and McMahon (1990) Biochem. J. 270, 591-597]. To further this observation, the cardiac membrane-bound form of arginine-specific mono-ADP-ribosyltransferase (EC 2.4.2.31) has been characterized. Apparent Km values of 330 and 470 microM were found in heart membrane preparations from rat and quail respectively. The Vmax. value depended greatly on the species of animal studied, and was 1.1 and 48 nmol/min per mg in rat and quail preparations respectively. The specific activity of the enzyme was lowest in pig, intermediate in rat, dog and rabbit, and highest in mouse and quail cardiac membranes. In the rat, the ADP-ribosylation of protein and enzyme activity were very low in heart preparations from 1-15-day-old animals. Thereafter the ADP-ribosylation and enzyme activity increased gradually to adulthood. Bacillus cereus phosphatidylinositol-specific phospholipase C, known to hydrolyse glycosylphosphatidylinositol anchors of proteins, released the mono-ADP-ribosyltransferase from membrane preparations of both rat and quail in a dose-dependent, Zn(2+)-inhibited manner. Thus it appears that a membrane-bound form of arginine-specific mono-ADP-ribosyltransferase is present in heart membranes from a variety of species and is not species-specific. The activity of this ADP-ribosyltransferase appears to be developmentally regulated and to be bound to the cardiac membranes by a glycosylphosphatidylinositol anchor.

Localization and partial characterization of ADP-ribosylation products in hearts from adult and neonatal rats.

The subcellular distributions of endogenous ADP-ribosylation products in hearts from 1-day-old neonatal and adult rats were investigated. In adult rat heart a 52 kDa mono-ADP-ribosylation product was identified in the plasma membrane fraction. In contrast, in neonatal rat heart a 130 kDa poly-ADP-ribosylation product was present in the nuclear fraction. The monomeric and polymeric nature of the two ADP-ribosylation products was determined by their sensitivity to thymidine and by analysis of their snake venom phosphodiesterase products. NADP+ enhanced both the mono- and polymeric reactions. The ADP-ribose-protein linkage of the adult 52 kDa product was stable to 1 h of treatment with hydroxylamine (0.5 M) and mercury ions, but was sensitive to alkali and a 12 h treatment with hydroxylamine (1 M). This is suggestive of an arginine linkage. The 130 kDa poly-ADP-ribosylation product from the neonatal rat heart was alkalilabile but stable to both hydroxylamine and HgCl2. This implies the presence of an unusual linkage in the 130 kDa product. The presence of these different ADP-ribosylation products in adult and neonatal rat hearts suggests the possible importance of these proteins and their ADP-ribosylation during cardiac development.