Isolation of a cDNA clone for human NAD+: protein ADP-ribosyltransferase.
NAD+:Protein ADP-ribosyltransferase (EC 2.4.2.30) (ADPRT) was purified from human placenta by affinity chromatography. With the purified enzyme specific antibodies were raised and partial amino acid sequences were determined. To one of the amino acid sequences corresponding oligonucleotides were synthesized. A sized HeLa lambda gt11 cDNA library was constructed and screened. Positive clones were characterized to be ADPRT specific by immuno- and hybridization techniques. Clone ADPRT-G8 reacted with affinity chromatographically purified specific antibodies and with two specific oligonucleotides. The DNA of this clone detected an mRNA of about 4 kb, sufficient in size to code for the ADPRT with an Mr of 116,000. Partial sequence analysis of this clone confirmed its identity by revealing sequences which code for peptides which were found in cyanogen bromide (CNBr) fragments of the purified enzyme. The ADPRT-G8 clone was characterized with respect to its restriction pattern. The cloned ADPRT cDNA now opens the possibility to investigate the role of this enzyme in control of cellular functions.
D036002:ADP Ribose Transferases; D000595:Amino Acid Sequence; D001483:Base Sequence; D002846:Chromatography, Affinity; D002851:Chromatography, High Pressure Liquid; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics*; D005260:Female; D006367:HeLa Cells; D006801:Humans; D007118:Immunoassay; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization; D009841:Oligonucleotides / Q000096:biosynthesis; D010430:Pentosyltransferases / Q000235:genetics* / Q000302:isolation & purification; D010449:Peptide Mapping; D010920:Placenta / Q000201:enzymology*; D011247:Pregnancy; D012333:RNA, Messenger / Q000032:analysis; D011993:Recombinant Fusion Proteins / Q000032:analysis
D009841:Oligonucleotides; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D004247:DNA; D036002:ADP Ribose Transferases; D010430:Pentosyltransferases