ADP-ribosylation of transducin by islet-activation protein. Identification of asparagine as the site of ADP-ribosylation.
Islet-activating protein catalyzes the ADP-ribosylation of transducin, a guanine nucleotide-binding regulatory protein that mediates activation of a retinal cyclic GMP-selective phosphodiesterase. Radiolabel from [adenylate-32P]NAD+ was incorporated specifically into the alpha subunit of purified transducin. Maximal levels of incorporation approximated 0.8 mol of ADP-ribose/mol of transducin. A peptide containing the ADP-ribosyl moiety was purified from a tryptic digest of radiolabeled transducin. This peptide was characterized by chemical and enzymatic procedures and by fast atom bombardment mass spectrometry. The primary structure of this peptide was Glu-Asn-Leu-Lys-Asn(ADP-ribose)-Gly-Leu-Phe. It is probable that the peptide originated from the carboxyl terminus of the alpha subunit and that the ADP-ribosyl moiety is attached by an N-glycosidic linkage to the asparagine residue. Transducin associated with retinal disc membranes is also ADP-ribosylated by cholera toxin. Cholera toxin and islet-activating protein sequentially catalyze the incorporation of 1.9 mol of ADP-ribose/mol of transducin, indicating two distinct sites of ADP-ribosylation within transducin.
D000246:Adenosine Diphosphate Ribose / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D001216:Asparagine / Q000378:metabolism*; D001426:Bacterial Proteins / Q000494:pharmacology*; D002268:Carboxypeptidases / Q000378:metabolism; D002417:Cattle; D002772:Cholera Toxin / Q000494:pharmacology; D013058:Mass Spectrometry; D008565:Membrane Proteins / Q000378:metabolism*; D009243:NAD / Q000378:metabolism; D009702:Nucleoside Diphosphate Sugars / Q000378:metabolism*; D037342:Pertussis Toxin; D012374:Rod Cell Outer Segment / Q000378:metabolism; D015293:Transducin; D010566:Virulence Factors, Bordetella
D001426:Bacterial Proteins; D008565:Membrane Proteins; D009702:Nucleoside Diphosphate Sugars; D010566:Virulence Factors, Bordetella; D009243:NAD; D000246:Adenosine Diphosphate Ribose; D001216:Asparagine; D002772:Cholera Toxin; D037342:Pertussis Toxin; D002268:Carboxypeptidases; D015293:Transducin