Biochemical Studies of Worm Insulins

描述(由申请人提供)： 秀丽线虫体内与胰岛素相关的信号通路控制着衰老和寿命。值得注意的是，线虫基因组包含38个假定的胰岛素样基因和一个人类胰岛素受体的同源基因(命名为daf-2)。线虫类胰岛素蛋白在结构和功能上真的与哺乳动物的胰岛素相似吗？为什么有这么多不同的配体？我们建议合成具有代表性的胰岛素样多肽(INS蛋白)，并研究它们的结构和受体结合特性。这项拟议的研究将阐明胰岛素结构的进化，并剖析衰老遗传模型中的信号通路。 拟议的研究集中在四个问题上。(1)考虑到它们的序列差异，Ins蛋白真的是胰岛素超家族的成员吗？如果是，不同的侧链是如何在保守的结构中调节的？(2)DAF-2是作为受体酪氨酸激酶发挥作用，如果是，它的激酶活性是受INS配体正向还是负向调节？(3)INS蛋白是否与人胰岛素受体结合，如果是，其看似不同的表面如何模仿人胰岛素？(4)相反，人胰岛素是否与DAF-2结合并调节？建议与盖伊·G·多德森教授进行长期合作，以比较激素受体复合体的共晶结构。对一种具有代表性的INS蛋白质的初步研究已经证实，尽管总体上有一个类似胰岛素的折叠，但其假定的受体结合表面与人类胰岛素的表面有很大的不同。看似自相矛盾的是，这种蛋白质仍然与人类的胰岛素受体结合并激活。人胰岛素同样与DAF-2受体结合。新型INS配体可能为设计人类胰岛素激动剂提供了一个意想不到的起点。考虑到胰岛素类似物对优化糖尿病治疗的日益重要，以及IGF-I拮抗剂在癌症治疗中可能的临床效用，对INS家族的研究可能在治疗蛋白质设计中具有翻译意义。

DESCRIPTION (provided by applicant): An insulin-related signaling pathway in the nematode Caenorhabditis elegans controls aging and life span. Remarkably, the C. elegans genome contains 38 putative insulin-like genes and a single orthologue of the human insulin receptor (designated daf-2). Do nematode insulin-like proteins truly resemble mammalian insulins in structure and function? Why are there such a diversity of ligands? We propose to synthesize representative insulin-like polypeptides (INS proteins) and investigate their structures and receptor-binding properties. The proposed studies will illuminate the evolution of the insulin structure and dissect a signaling pathway in a genetic model of senescence. The proposed studies focus on four questions. (1) Given their sequence divergence, are INS proteins truly members of the insulin superfamily, and if so, how are the divergent side chains accommodated within a conserved architecture? (2) Does DAF-2 function as a receptor tyrosine kinase, and if so, is its kinase activity positively or negatively regulated by INS ligands? (3) Do INS proteins bind to the human insulin receptor, and if so, how can their seemingly divergent surfaces mimic human insulin? (4) Conversely, does human insulin bind to and regulate DAF-2? A long-term collaboration is proposed with Prof. Guy G. Dodson to compare co-crystal structures of hormone-receptor complexes. Preliminary studies of a representative INS protein have established that -- despite an overall insulin-like fold -- its putative receptor-binding surface differs dramatically from that of human insulin. As a seeming paradox, the protein nonetheless binds to and activates the human insulin receptor. Human insulin likewise binds to the DAF-2 receptor. Novel INS ligands may provide an unexpected starting point for design of human insulin agonists. Given the growing importance of insulin analogs to optimize treatment of diabetes mellitus and the possible clinical utility of IGF-I antagonists in cancer therapy, studies of the INS family may have translational implications in therapeutic protein design.