PHOSPHOPEPTIDE MODELS OF P53 PROTEIN FRAGMENTS

描述：野生型P53蛋白在调节 生长、G1/S边界停滞、反式激活和细胞凋亡。基因突变 在一半的人类肿瘤中发现了p53基因。循环自身抗体 在10%-15%的癌症患者中发现了P53。虽然在场的 的自身抗体与p53突变重合，这些自身抗体是 不是特定的p53突变体所特有的，并且可以识别野生型和 突变型蛋白表明P53的免疫原性不是 突变的直接后果。P53是一种磷蛋白，含有 大量建议的或潜在的磷酸化位点。 此外，还鉴定了一个或多个O-连接的糖基化位点。 尽管有广泛的研究将磷酸化与肿瘤联系起来 抑制或缺乏抑制，知道哪些磷酸盐受体位点是 磷酸化的终极生物学意义是非常重要的 有限的。以前对突变蛋白质的研究一直很复杂，因为 在使用的细胞系中的磷酸化机制中的潜在异常 用于蛋白质生产。这项提议的目标是明确地 鉴定野生型和突变型的磷酸化和糖基化位点 P53。为此，很好地表征了单和多个磷酸化和 糖基化多肽将被合成并用作小鼠的免疫原 制备高滴度、高特异性的单抗 抗原的修饰形式。将对抗体进行抗原检测 对广泛的p53等位基因的识别。合成的多肽将 是筛查癌症P53自身抗体的有价值的工具 患者判断自身抗体是简单识别还是多识别 抗原结构的复杂修饰。在确定了实际的 正常细胞和癌细胞中的磷酸化位点未来的研究可能 目标(1)激活或解除对适当的激活酶和 磷酸酶和(2)免疫和免疫治疗的设计方案 针对修饰后的P53。

DESCRIPTION: The wild-type p53 protein plays a central role in regulating growth, G1/S boundary arrest, transactivation and apoptosis. Mutations in the p53 gene are found in half of human tumors. Circulating autoantibodies to p53 are found in 10-15 percent of cancer patients. Although the presence of p53 autoantibodies coincides with p53 mutations, these autoantibodies are not specific to a particular p53 mutant and recognize both wild type and mutant proteins therefore indicating that the immunogenicity of p53 is not a direct consequence of the mutations. p53 is a phosphoprotein that contains a large number of proposed or potential phosphorylation sites. Additionally, one or more O-linked glycosylation sites have been identified. In spite of extensive research to correlate phosphorylation and tumor suppression or lack thereof, knowledge of which phosphate acceptor sites are utilized and the ultimate biological significance of phosphorylation is very limited. Previous studies on mutated proteins have been complicated because of potential anomalies in the phosphorylation machinery in cell lines used for protein production. The goal of the proposal is to unambiguously identify phosphorylation and glycosylation sites in wild type and mutant p53. To this end well-characterized single and multiply phosphorylated and glycosylated peptides will be synthesized and used as immunogens in mice to generate monoclonal antibodies with high titer and specificity for the modified forms of the antigens. The antibodies will be tested for antigen recognition of a wide range of p53 alleles. The synthetic peptides will serve as valuable tools for screening p53 autoantibodies from cancer patients to determine whether the autoantibodies recognize simple or more complex modifications in antigen structure. After identifying the actual phosphorylation sites in normal and cancerous cells future research may target (i) the activation or deregulation of appropriate kinases and phosphatases and (ii) design protocols for immunization and immunotherapy against modified p53.