title: Comparative biophysical characterization of p53 with the pro-apoptotic BAK and the anti-apoptotic BCL-xL.
abstract: The p53 transcription-independent apoptosis in mitochondria, mediated by its interaction with the pro-apoptotic and the anti-apoptotic members of the Bcl2 family of proteins, has been described in vivo, especially in radiosensitive tissues. We have characterized the interaction of p53 with both the pro-apoptotic Bak and the anti-apoptotic Bcl-x(L) proteins, comparing their affinity and their interaction surfaces, using biophysical techniques such as fluorescence anisotropy, analytical ultracentrifugation, and NMR. We have shown that both proteins interact with only the p53 core domain and not with its N- and C-terminal regions. Further, p53 has a higher affinity for Bcl-x(L) than for Bak, which is consistent with the previously described sequential binding of Bcl-x(L) and Bak by p53. Interestingly, although the interaction with both proteins is electrostatic in character, they have different binding sites. Using NMR spectroscopy, we have determined that Bcl-x(L) interacts with the DNA binding site of p53, but Bak does not interact with this site. A new potential interaction surface for Bak is proposed.
authors: Medical Research Council, Centre for Protein Engineering, Hills Road, Cambridge CB2 0QH, United Kingdom.
authors: Begona
authors: B
authors: Sot
authors: Stefan M V
authors: SM
authors: Freund
authors: Alan R
authors: AR
authors: Fersht
mesh_terms: D000818:Animals; D016880:Anisotropy; D001665:Binding Sites; D001667:Binding, Competitive; D006801:Humans; D009682:Magnetic Resonance Spectroscopy; D008856:Microscopy, Fluorescence; D008954:Models, Biological; D008968:Molecular Conformation; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism*; D014461:Ultracentrifugation; D050998:bcl-2 Homologous Antagonist-Killer Protein / Q000502:physiology*; D051020:bcl-X Protein / Q000378:metabolism*
chemical_list: C494772:BAK1 protein, human; D016159:Tumor Suppressor Protein p53; D050998:bcl-2 Homologous Antagonist-Killer Protein; D051020:bcl-X Protein