title: ADP-ribosylation of wild-type p53 in vitro: binding of p53 protein to specific p53 consensus sequence prevents its modification.
abstract: We have recently reported that mutant but not wild-type (wt) p53 protein was ADP-ribosylated in primary rat cells overexpressing the temperature-sensitive murine p53val135 gene. To examine whether the lack of susceptibility to modification is a specific feature of p53val135 adopting wt conformation or rather a general property of this tumor suppressor protein, we have studied ADP-ribosylation of wt p53 of different origin in vitro using semi-purified poly(ADP-ribose) transferase (pADPRT). In vitro pADPRT modified human and mouse wt p53 and p53val135. Under limiting substrate concentration, the molar mass of ADP-ribosylated p53 was only slightly altered. Chase experiments with high NAD concentration resulted in the formation of poly(ADP-ribosyl)ated p53 protein shifted to 64 kD. However, preincubation of wt p53 proteins with a p53 consensus sequence resulting in complex formation abolished the modification of wt p53. This indicates that in the cellular environment the specific DNA binding of wt p53 prevents its covalent modification by poly(ADP-ribose).
authors.affiliation: Institute of Tumorbiology-Cancer Research, University of Vienna, Austria.
authors.forename: J
authors.initials: J
authors.lastname: Wesierska-Gadek
authors.forename: G
authors.initials: G
authors.lastname: Schmid
authors.forename: C
authors.initials: C
authors.lastname: Cerni
mesh_terms: D000246:Adenosine Diphosphate Ribose / Q000378:metabolism; D000818:Animals; D002478:Cells, Cultured; D016384:Consensus Sequence; D004622:Embryo, Mammalian; D004625:Embryo, Nonmammalian; D006801:Humans; D051379:Mice; D009243:NAD / Q000378:metabolism; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism*; D051381:Rats; D011916:Rats, Inbred F344; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000378:metabolism; D018411:Spodoptera; D013379:Substrate Specificity; D014162:Transfection; D016159:Tumor Suppressor Protein p53 / Q000096:biosynthesis / Q000378:metabolism*
chemical_list: D011994:Recombinant Proteins; D016159:Tumor Suppressor Protein p53; D009243:NAD; D000246:Adenosine Diphosphate Ribose; D011065:Poly(ADP-ribose) Polymerases