EFFECT OF PHOSPHORYLATION ON TETRAMERIZATION OF THE TUMOR-SUPPRESSOR PROTEIN P53

Human tumor suppresses protein p53 is a 393-amino acid phosphoprotein that enhances transcription in response to DNA damage from several gen es that regulate cell cycle progression. The tetrameric state of p53 i s critical to wild-type function; the p53 tetramerization element is l ocated in the C-terminal region of the protein. This region is phospho rylated at several evolutionarily conserved serines, suggesting that p hosphorylation may be an important regulator of p53 function. In order to determine the effect of phosphorylation on tetramer formation, we synthesized phosphopeptides corresponding to p53(Ser303-Asp393) with p hosphate incorporated at Ser315, Ser378, or Ser392, and at both Ser315 and Ser392. Equilibrium ultracentrifugation analysis showed that phos phorylation at Ser392 increased the association constant for tetramer formation nearly ten-fold. By itself, phosphorylation at Ser315 or Ser 378 had little effect on tetramer formation, but Ser315 largely revers ed the effect of phosphorylation at Ser392. Analysis by calorimetry su ggests that phosphorylation may influence subunit affinity by an entha lpy driven process.