ADP-ribosyltransferase in isolated nuclei from sea-urchin embryos.
The activity of ADP-ribosyltransferase in nuclei isolated from sea-urchin embryos was estimated by the incorporation of [adenosine-14C]NAD+ into the acid-insoluble fraction. Hydrolysis of this acid-insoluble product by snake venom phosphodiesterase yielded radioactive 5'-AMP and phosphoribosyl-AMP. The incorporation of [14C]-NAD+ was inhibited by 3-aminobenzamide and nicotinamide, potent inhibitors of ADP-ribosyltransferase. [14C]NAD+ incorporation into the acid-insoluble fraction results from the reaction of ADP-ribosyltransferase. The optimum pH for the enzyme in isolated nuclei was 7.5. The enzyme, in 50 mM-Tris/HCl buffer, pH 7.5, containing 0.5 mM-NAD+ and 0.5 mM-dithiothreitol, exhibited the highest activity at 18 degrees C in the presence of 14 mM-MgCl2. The apparent Km value for NAD+ was 25 microM. The activity of the enzyme was measured in nuclei isolated from the embryos at several stages during early development. The activity was maximum at the 16-32-cell stage and then decreased to a minimum at the mesenchyme blastula stage. Thereafter its activity slightly increased at the onset of gastrulation and decreased again at the prism stage.
Isoai
A
A
Yasumasu
I
I
D000818:Animals; D002467:Cell Nucleus / Q000187:drug effects / Q000201:enzymology; D006863:Hydrogen-Ion Concentration; D008274:Magnesium / Q000494:pharmacology; D009713:Nucleotidyltransferases / Q000378:metabolism*; D011064:Poly Adenosine Diphosphate Ribose / Q000096:biosynthesis; D011065:Poly(ADP-ribose) Polymerases; D012617:Sea Urchins / Q000187:drug effects / Q000196:embryology / Q000201:enzymology*; D013696:Temperature; D013997:Time Factors
D011064:Poly Adenosine Diphosphate Ribose; D011065:Poly(ADP-ribose) Polymerases; D009713:Nucleotidyltransferases; D008274:Magnesium