4 P53 DNA-BINDING DOMAIN PEPTIDES BIND NATURAL P53-RESPONSE ELEMENTS AND BEND THE DNA
4 P53 DNA-BINDING DOMAIN PEPTIDES BIND NATURAL P53-RESPONSE ELEMENTS AND BEND THE DNA
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DOI：
10.1073/pnas.92.19.8591
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发表时间：
1995-09-12
影响因子：
11.1
通讯作者：
HARRINGTON, RE
中科院分区：
文献类型：
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作者：
BALAGURUMOORTHY, P;SAKAMOTO, H;HARRINGTON, RE
最近对p53的最小核心DNA结合域(P53DBD)的结构研究已经为它们的功能提供了有价值的见解，但关于它们的相互作用和协同作用的许多问题仍然没有得到回答。为了更好地理解这些关系，我们用DNA环化和分析超速离心法研究了p53DBD与两个具有重要生物学意义的全反应元件(WAF1和核糖体基因簇位点)的结合，我们发现p53DBD与DNA强烈结合，并协同作用于p53DBD和p53DBD与DNA的结合计量比为4：1。平均表观K-d为(8.3+/-1.4)×10(-8)M，没有检测到低化学计量比的中间物种，我们进一步表明，络合物的形成在两个响应元件中都诱导了至少60度的轴向弯曲。综上所述，这些结果表明，p53DBD具有指导形成与野生型p53相同的4：1 DNA结合化学计量比的紧密核蛋白复合体的能力，并伴随着反应元件DNA的显著构象变化，这表明p53DBD可能在P53的四聚功能中发挥作用，并提出了这方面的可能作用。
Recent structural studies of the minimal core DNA-binding domain of p53 (p53DBD) complexed to a single consensus pentamer sequence and of the isolated p53 tetramerization domain have provided valuable insights into their functions, but many questions about their interacting roles and synergism remain unanswered, To better understand these relationships, we have examined the binding of the p53DBD to two biologically important full-response elements (the WAF1 and ribosomal gene cluster sites) by using DNA circularization and analytical ultracentrifugation, We show that the p53DBD binds DNA strongly and cooperatively with p53DBD to DNA binding stoichiometries of 4:1. For the WAF1 element, the mean apparent K-d is (8.3 +/- 1.4) x 10(-8) M, and no intermediate species of lower stoichiometries can be detected, We show further that complex formation induces an axial bend of at least 60 degrees in both response elements. These results, taken collectively, demonstrate that p53DBD possesses the ability to direct the formation of a tight nucleoprotein complex having the same 4:1 DNA-binding stoichiometry as wild-type p53 which is accompanied by a substantial conformational change in the response-element DNA, This suggests that the p53DBD may play a role in the tetramerization function of p53, A possible role in this regard is proposed.