The induction of apoptosis by p53 in response to cellular stress is its most conserved function and crucial for p53 tumor suppression. We recently reported that p53 directly induces oligomerization of the BH1,2,3 effector protein Bak, leading to outer mitochondrial membrane permeabilization (OMMP) with release of apoptotic activator proteins. One important mechanism by which p53 achieves OMMP is by forming an inhibitory complex with the antiapoptotic BclXL protein. In contrast, the p53 complex with the Bcl2 homolog has not been interrogated. Here we have undertaken a detailed characterization of the p53-Bcl2 interaction using structural, biophysical, and mutational analyses. We have identified the p53 DNA binding domain as the binding interface for Bcl2 using solution NMR. The affinity of the p53-Bcl2 complex was determined by surface plasmon resonance analysis (BIAcore) to have a dominant component K D 535 ؎ 24 nM. Moreover, in contrast to wild type p53, endogenous missense mutants of p53 are unable to form complexes with endogenous Bcl2 in human cancer cells. Functionally, these mutants are all completely or strongly compromised in mediating OMMP, as measured by cytochrome c release from isolated mitochondria. These data implicate p53-Bcl2 complexes in contributing to the direct mitochondrial p53 pathway of apoptosis and further support the notion that the DNA binding domain of p53 is a dual function domain, mediating both its transactivation function and its direct mitochondrial apoptotic function.A major function of the p53 tumor suppressor is the induction of an apoptotic program in response to a broad variety of cell stresses. Thus, understanding the mechanisms by which p53 executes cell death pathways is of considerable importance in cancer biology. The basis for the powerful apoptotic and tumor suppressor activity of p53 lies in its pleiotropism, which includes transcription-dependent and transcription-independent functions (1, 2). p53-mediated apoptosis primarily signals through the mitochondrial pathway (1). Some notable p53 target genes such as the BH3-only proteins PUMA, Noxa, Bax, and p53AIP1 reside and/or act at the mitochondria (3-7).We previously showed that in response to a death stimulus such as DNA damage or hypoxia, a fraction of stabilized p53 rapidly translocates to mitochondria in primary, immortal, and transformed cells (8 -11). The functional consequences of this phenomenon were revealed by targeting exogenous p53 to mitochondria in p53 null cells. Mitochondrially targeted p53 was sufficient to launch apoptosis and suppress colony formation directly from the mitochondrial platform in a transcription-independent fashion (9, 10). Translocated endogenous mitochondrial p53 interacts with anti-apoptotic BclXL and Bcl2 proteins and blocks their functions. Purified p53 protein induces oligomerization of Bak and permeabilization of the outer mitochondrial membrane and strongly promotes cytochrome c release from healthy unstressed mitochondria (10).Using computational and mutational analyses, we ...

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Mutations in the transcription factor p53 enhance one's predisposition to cancer. Mutational studies show that double salt bridges are key elements for the dimerization of p53, its cooperative binding to DNA, and thus its proper function as a tumor suppressor (see picture). This might explain certain germ‐line mutations found in patients with Li–Fraumeni syndrome.

Every single day, the DNA of each cell in the human body is mutated thousands of times, even in absence of oncogenes or extreme radiation. Many of these mutations could lead to cancer and, finally, death. To fight this, multicellular organisms have evolved an efficient control system with the tumor-suppressor protein p53 as the central element. An intact p53 network ensures that DNA damage is detected early on. The importance of p53 for preventing cancer is highlighted by the fact that p53 is inactivated in more than 50 % of all human tumors. Thus, for good reason, p53 is one of the most intensively studied proteins. Despite the great effort that has been made to characterize this protein, the complex function and the structural properties of p53 are still only partially known. This review highlights basic concepts and recent progress in understanding the structure and regulation of p53, focusing on emerging new mechanistic and therapeutic concepts.

Hsp90 is one of the most abundant chaperone proteins in the cytosol. In an ATP-dependent manner it plays an essential role in the folding and activation of a range of client proteins involved in signal transduction and cell cycle regulation. We used NMR shift perturbation experiments to obtain information on the structural implications of the binding of AMP-PNP (adenylyl-imidodiphosphate-a non-hydrolysable ATP analogue), ADP and the inhibitors radicicol and geldanamycin. Analysis of (1)H,(15)N correlation spectra showed a specific pattern of chemical shift perturbations at N210 (ATP binding domain of Hsp90, residues 1-210) upon ligand binding. This can be interpreted qualitatively either as a consequence of direct ligand interactions or of ligand-induced conformational changes within the protein. All ligands show specific interactions in the binding site, which is known from the crystal structure of the N-terminal domain of Hsp90. For AMP-PNP and ADP, additional shift perturbations of residues outside the binding pocket were observed and can be regarded as a result of conformational rearrangement upon binding. According to the crystal structures, these regions are the first alpha-helix and the "ATP-lid" ranging from amino acids 85 to 110. The N-terminal domain is therefore not a passive nucleotide-binding site, as suggested by X-ray crystallography, but responds to the binding of ATP in a dynamic way with specific structural changes required for the progression of the ATPase cycle.