Mechanistic insights into p53-promoted RNA-RNA annealing.
The tumour suppressor protein p53 promotes the annealing of complementary nucleic acids in vitro. We observed an up to 1600-fold increase of RNA-RNA annealing by recombinant p53 protein which was shown to bind to RNA in sequence-independent way. Nuclease mapping experiments suggest that p53 binds to intramolecular duplex portions and only marginally changes the overall secondary structure of RNA at conditions of increased annealing. Thus, the mechanism of p53-promoted RNA-RNA annealing does not seem to be dependent on an activity that melts or changes RNA structure. The activation enthalpy of RNA-RNA annealing is decreased in the presence of p53, i.e. the p53 protein could stabilize the transition state whereas the activation entropy is unfavourable. A comparison with thermodynamic data measured for other facilitators strongly suggests that the mechanism of p53-promoted RNA-RNA annealing is distinct from the mechanism by which other facilitators work. The annealing activity of p53 is almost abolished in the presence of magnesium indicating that it can be sharply regulated in vitro and, in principle, could also be regulated in vivo.
Nedbal
W
W
Forschungsschwerpunkt Angewandte Tumorvirologie Deutsches Krebsforschungszentrum, Heidelberg, Germany.
Frey
M
M
Willemann
B
B
Zentgraf
H
H
Sczakiel
G
G
D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D009690:Nucleic Acid Conformation; D009691:Nucleic Acid Denaturation; D009693:Nucleic Acid Hybridization; D012313:RNA / Q000737:chemistry* / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D012260:Ribonucleases / Q000378:metabolism; D013696:Temperature; D013816:Thermodynamics; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism*
D011994:Recombinant Proteins; D016159:Tumor Suppressor Protein p53; D012313:RNA; D012260:Ribonucleases; D008274:Magnesium