Human nuclear NAD sup + ADP-ribosyltransferase: Localization of the gene on chromosome 1q41-q42 and expression of an active human enzyme in Escherichia coli

The gene for human nuclear NAD{sup +} ADP-ribosyltransferase was localized to chromosome 1 at q41-q42 by in situ hybridization with a pADPRT-specific cDNA probe. Expression of a pAD-PRT cDNA under control of the lac promoter in Escherichia coli induces the synthesis of a group of related proteins that were immunoreactive with pADPRT antibody and that had catalytic properties very similar to those of the human enzyme. Purification of this enzymatic activity was performed essentially as described for the human enzyme. The K{sub m}, pH optimum, optimal reaction temperature, and inhibition by 3-aminobenzamide and 3-methoxybenzamide were found to be similar for the recombinant and the human enzymes. The purified recombinant enzyme consists of two major proteins of M{sub r} 99,000 and M{sub r} 89,000. Both proteins show pADPRT activity in activity gel analysis with ({sup 32}P)NAD{sup +} as substrate. Microsequencing of these two proteins isolated by denaturing gel electrophoresis and deletion mutagenesis of the pADPRT expression plasmid shows that the M{sub r} 99,000 and M{sub r} 89,000 proteins derive from initiation of translation at interval translational start signals located within the pADPRT cDNA.