title: High thermostability and lack of cooperative DNA binding distinguish the p63 core domain from the homologous tumor suppressor p53.
abstract: The p53 protein is the major tumor suppressor in mammals. The discovery of the p53 homologs p63 and p73 defined a family of p53 members with distinct roles in tumor suppression, differentiation, and development. Here, we describe the biochemical characterization of the core DNA-binding domain of a human isoform of p63, p63-delta, and particularly novel features in comparison with p53. In contrast to p53, the free p63 core domain did not show specific binding to p53 DNA consensus sites. However, glutathione S-transferase-fused and thus dimerized p63 and p53 core domains had similar affinity and specificity for the p53 consensus sites p21, gadd45, cyclin G, and bax. Furthermore, the fold of p63 core was remarkably stable compared with p53 as judged by differential scanning calorimetry (T(m) = 61 degrees C versus 44 degrees C for p53) and equilibrium unfolding ([urea](50%) = 5.2 m versus 3.1 m for p53). A homology model of p63 core highlights differences at a segment near the H1 helix hypothetically involved in the formation of the dimerization interface in p53, which might reduce cooperativity of p63 core DNA binding compared with p53. The model also shows differences in the electrostatic and hydrophobic potentials of the domains relevant to folding stability.
authors.affiliation: Pharma Research, Roche Diagnostics GmbH, D-82372 Penzberg, Germany. christian.klein.ck1@roche.com
authors.forename: C
authors.initials: C
authors.lastname: Klein
authors.forename: G
authors.initials: G
authors.lastname: Georges
authors.forename: K P
authors.initials: KP
authors.lastname: Künkele
authors.forename: R
authors.initials: R
authors.lastname: Huber
authors.forename: R A
authors.initials: RA
authors.lastname: Engh
authors.forename: S
authors.initials: S
authors.lastname: Hansen
mesh_terms: D000595:Amino Acid Sequence; D002942:Circular Dichroism; D003001:Cloning, Molecular; D004247:DNA / Q000378:metabolism*; D004268:DNA-Binding Proteins; D016147:Genes, Tumor Suppressor; D006801:Humans; D008565:Membrane Proteins; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010750:Phosphoproteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011489:Protein Denaturation; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid; D013050:Spectrometry, Fluorescence; D013696:Temperature; D013816:Thermodynamics; D015534:Trans-Activators / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014157:Transcription Factors; D016159:Tumor Suppressor Protein p53 / Q000737:chemistry / Q000378:metabolism*; D025521:Tumor Suppressor Proteins; D014508:Urea / Q000494:pharmacology; D015032:Zinc / Q000378:metabolism
chemical_list: C081876:CKAP4 protein, human; D004268:DNA-Binding Proteins; D008565:Membrane Proteins; D010750:Phosphoproteins; C115051:TP63 protein, human; D015534:Trans-Activators; D014157:Transcription Factors; D016159:Tumor Suppressor Protein p53; D025521:Tumor Suppressor Proteins; D014508:Urea; D004247:DNA; D015032:Zinc