Sakaguchi K, Sakamoto H, Xie D, Erickson J W, Lewis M S, Anderson C W, Appella E
Laboratory of Cell Biology, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.
J Protein Chem. 1997 Jul;16(5):553-6. doi: 10.1023/a:1026334116189.
Human tumor suppressor protein p53 is a 393-amino acid phosphoprotein that enhances transcription in response to DNA damage from several genes that regulate cell cycle progression. The tetrameric state of p53 is critical to wild-type function; the p53 tetramerization element is located in the C-terminal region of the protein. This region is phosphorylated at several evolutionarily conserved serines, suggesting that phosphorylation 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 phosphate incorporated at Ser315, Ser378, or Ser392, and at both Ser315 and Ser392. Equilibrium ultracentrifugation analysis showed that phosphorylation at Ser392 increased the association constant for tetramer formation nearly ten-fold. By itself, phosphorylation at Ser315 or Ser378 had little effect on tetramer formation, but Ser315 largely reversed the effect of phosphorylation at Ser392. Analysis by calorimetry suggests that phosphorylation may influence subunit affinity by an enthalpy driven process.
人类肿瘤抑制蛋白p53是一种含393个氨基酸的磷蛋白，它可响应来自多个调控细胞周期进程的基因的DNA损伤，增强转录。p53的四聚体状态对野生型功能至关重要；p53四聚化元件位于该蛋白的C末端区域。该区域在几个进化保守的丝氨酸处被磷酸化，这表明磷酸化可能是p53功能的重要调节因子。为了确定磷酸化对四聚体形成的影响，我们合成了与p53(Ser303 - Asp393)相对应的磷酸肽，其中磷酸基团分别掺入Ser315、Ser378或Ser392，以及同时掺入Ser315和Ser392。平衡超速离心分析表明，Ser392处的磷酸化使四聚体形成的缔合常数增加了近10倍。单独来看，Ser315或Ser378处的磷酸化对四聚体形成影响不大，但Ser315在很大程度上逆转了Ser392处磷酸化的作用。量热分析表明，磷酸化可能通过一个焓驱动的过程影响亚基亲和力。
