The CBP/p300 TAZ1 domain in its native state is not a binding partner of MDM2.

The transcriptional co-activator CBP [CREB (cAMP-response-element-binding protein)-binding protein] and its paralogue p300 play a key role in the regulation of both activity and stability of the tumour suppressor p53. Degradation of p53 is mediated by the ubiquitin ligase MDM2 (mouse double minute protein) and is also reported to be regulated ...
by CBP/p300. Direct protein-protein interaction between a central domain of MDM2 and the TAZ1 (transcriptional adaptor zinc-binding domain) [C/H1 (cysteine/histidine-rich region 1)] domain of p300 and subsequent formation of a ternary complex including p53 have been reported previously. We expressed and purified the proposed binding domains of HDM2 (human homologue of MDM2) and CBP, and examined their interactions using CD spectroscopy. The binding studies were extended by using natively purified GST (glutathione S-transferase)-p300 TAZ1 and GST-p53 fusion proteins, together with in vitro translated HDM2 fragments, under similar solution conditions to those in previous studies, but omitting added EDTA, which causes unfolding and aggregation of the zinc-binding TAZ1 domain. Comparing the binding properties of the known TAZ1 interaction partners HIF-1alpha (hypoxia-inducible factor 1), CITED2 (CBP/p300-interacting transactivator with glutamic- and aspartic-rich tail) and STAT2 (signal transducer and activator of transcription 2) with HDM2, our data suggest that TAZ1 in its native state does not serve as a specific recognition domain of HDM2. Rather, unfolded TAZ1 and HDM2 proteins have a high tendency to aggregate, and non-specific protein complexes are formed under certain conditions.
Mesh Terms:
Animals, CREB-Binding Protein, Circular Dichroism, Cloning, Molecular, E1A-Associated p300 Protein, Escherichia coli, Glutathione Transferase, Humans, Mice, Nuclear Proteins, Peptide Fragments, Peptides, Protein Binding, Protein Interaction Mapping, Protein Structure, Tertiary, Proto-Oncogene Proteins, Proto-Oncogene Proteins c-mdm2, Recombinant Fusion Proteins, Trans-Activators, Transfection, Zinc Fingers
Biochem. J.
Date: Aug. 01, 2004
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