A stable human p53 heterotetramer based on constructive charge interactions within the tetramerization domain.
The human p53 tetramerization domain (called p53tet; residues 325-355) spontaneously forms a dimer of dimers in solution. Hydrophobic interactions play a major role in stabilizing the p53 tetramer. However, the distinctive arrangement of charged residues at the dimer-dimer interface suggests that they also contribute to tetramer stability. Charge-reversal mutations at positions 343, 346, and 351 within the dimer-dimer interface were thus introduced into p53tet constructs and shown to result in the selective formation of a stable heterotetramer composed of homodimers. More precisely, mutants p53tet-E343K/E346K and p53tet-K351E preferentially associated with each other, but not with wild-type p53tet, to form a heterodimeric tetramer with enhanced thermal stability relative to either of the two components in isolation. The p53tet-E343K/E346K mutant alone assembled into a weakly stable tetramer in solution, whereas p53tet-K351E existed only as a dimer. Moreover, these mutants did not form heterocomplexes with wild-type p53tet, illustrating the specificity of the ionic interactions that form the novel heterotetramer. This study demonstrates the dramatic importance of ionic interactions in altering the stability of the p53 tetramer and in selectively creating heterotetramers of this protein scaffold.
Brokx
Richard D
RD
Department of Medical Biophysics, University of Toronto, Ontario M5G 2M9, Canada.
Bolewska-Pedyczak
Eleonora
E
Gariépy
Jean
J
D000595:Amino Acid Sequence; D002942:Circular Dichroism; D019281:Dimerization; D004591:Electrophoresis, Polyacrylamide Gel; D014644:Genetic Variation; D006801:Humans; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D013696:Temperature; D013997:Time Factors; D016159:Tumor Suppressor Protein p53 / Q000737:chemistry* / Q000378:metabolism*; D014461:Ultracentrifugation
D016159:Tumor Suppressor Protein p53