The N terminus of p53 regulates its dissociation from DNA

It is important to gain insight into p53 DNA binding and how it is regulate d. By using electrophoretic mobility shift assays and DNase I footprinting, we show that a region within the N terminus of the protein controls the di ssociation of p53 from a p53-binding site. When p53 is bound by a number of N-terminal-specific monoclonal antibodies, its rate of dissociation from D NA is reduced, and its ability to protect a cognate site from DNase I diges tion is increased. Moreover, greatly reduced dissociation is observed with p53 protein lacking the N-terminal 96 amino acids. By contrast, deletion of the C terminus does not affect p53 dissociation from DNA or DNase I protec tion. p53 protein expressed in and purified from bacterial cells displays m arkedly more instability on its consensus DNA-binding site than does p53 pr oduced in insect cells, suggesting that post-translational modifications ma y affect the stability of the protein. Our results provide evidence that th e N terminus of p53 possesses an auto-inhibitory function that is mechanist ically different from the inhibitory region at the C terminus.