DISTINCT RESIDUES OF HUMAN P53 IMPLICATED IN BINDING TO DNA, SIMIAN-VIRUS-40 LARGE T-ANTIGEN, 53BP1, AND 53BP2

We identified a minimal domain of human p53 required for the transacti vation of a p53 response element in Saccharomyces cerevisiae. This dom ain contains the central region of p53 sufficient for specific DNA bin ding, which colocalizes with the region responsible for binding simian virus 40 large T antigen, 53BP1, and 53BP2. Thirty amino acid positio ns, including natural mutational hot spots (R175, R213, R248, R249, an d R273), in the minimal DNA-binding domain were mutated by alanine sub stitution. Alanine substitutions at positions R213, R248, R249, D281, R282, R283, E286, and N288 affected transactivation but allowed bindin g to at least one of the three interacting proteins; these amino acids may be involved in amino acid-base pair contacts. Surprisingly, alani ne substitution at the mutational hot spot R175 did not affect DNA bin ding, transactivation, or T-antigen binding, although it nearly elimin ated binding to 53BP1 and 53BP2. Mutation of H168 significantly affect ed only T-antigen binding, and mutation of E285 affected only 53BP1 bi nding. Thus, we implicate specific residues of p53 in different DNA an d protein interactions.