ADP-ribosyltransferase is highly conserved: purification and characterization of ADP-ribosyltransferase from a fish and its comparison with the human enzyme.

Covalent modification of proteins by ADP-ribosylation is a major mode of protein regulation in eukaryotic cells. ADP-ribosyltransferases have been characterized from mammals but little is known about these enzymes in lower vertebrates. We purified an ADP-ribosyltransferase (E.C. 2.4.2.30) from trout (Salmo trutta faris) by affinity chromatography and characterized it. The 11,700-fold purified activity shows a major protein band at a molecular mass of 75,000 kDa in a SDS-polyacrylamide gel. In situ reactivation of SDS gels showed the 75,000 kDa protein to be enzymatically active, and additional enzymatically active bands at molecular masses of 115,000, 90,000 and 87,000 kDa, respectively. The enzyme is capable of poly-ADP-ribosylation. It crossreacts with affinity purified antibodies raised against human poly(ADP-ribose)synthetase and, except for the temperature optimum, its properties strongly resemble the mammalian enzymes, indicating the conserved character of nuclear ADP-ribosyltransferases. The trout enzyme is DNA- and histone-dependent, has an optimal pH between 8 and 9 and an apparent Km for NAD+ of 24 microM. The temperature optimum is 10 degrees C compared with 25 degrees C for the human enzyme. Known ADP-ribosyltransferase inhibitors also inhibit the enzyme from trout.