3D STRUCTURE OF THE CYCLIC AMP DEPENDENT PROTEIN KINASE

环放大器依赖性蛋白激酶的 3D 结构

• 批准号: 2182762
• 负责人: Jeff Kuret
• 金额: $ 3.63万
• 依托单位: APPLIED NEUROSOLUTIONS, INC.
• 依托单位国家: 美国
• 财政年份: 1991
• 资助国家: 美国
• 起止时间: 1991-05-01 至 1995-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2182762
• 关键词: Saccharomyces cerevisiae; X ray crystallography; active sites; chimeric proteins; computer simulation; conformation; enzyme inhibitors; enzyme structure; enzyme substrate; enzyme substrate analog; enzyme substrate complex; fungal genetics; isomorphous substitution; isozymes; molecular dynamics; physical model; protein engineering; protein folding; protein kinase; protein kinase A

Protein kinases are key regulatory molecules that participate in many biochemical pathways. As catalysts, these enzymes can amplify intra- or extracellular signals. By virtue of their broad yet limited substrate specificity, they are able to regulate more than one substrate protein at a time, coordinating intracellular metabolism. Understanding how protein kinases achieve the specificity necessary to regulate cell processes is an important problem in biology, and is the long-term goal of this proposal. Specifically, a detailed structural study of TPK1, the cAMP-dependent protein kinase catalytic subunit of Saccharomyces cerevisiae, is proposed. Existing crystals of TPK1 that diffract to 2.2 A resolution will be derivatized with heavy atom compounds, solving the crystallographic phase problem, and yielding its three-dimensional structure. The structure will reveal the location of key residues previously demonstrated to be involved in substrate recognition and catalysis, and reveal the overall folding of a protein kinase domain. This work will be extended by predicting the structure of other protein kinases that differ from TPK1 in substrate specificity using comparative modeling methods. The resulting prediction of protein kinase substrate selectively will be tested by in vitro mutagenesis. Chimeric kinases will be synthesized and characterized to relate the structural features predicted by computer modeling to enzymatic activity. A complete understanding of protein kinase structure will speed the development of specific inhibitors potentially useful in the treatment of neoplastic disease.

蛋白激酶是关键的调节分子，参与许多 生化途径 作为催化剂，这些酶可以放大细胞内或细胞外的 细胞外信号 凭借其广泛而有限的基底 特异性，它们能够调节一个以上的底物蛋白在一个 时间，协调细胞内代谢。 了解蛋白质如何 激酶实现调节细胞过程所必需的特异性是一种 这是生物学中的一个重要问题，也是本提案的长期目标。 具体地说，详细的结构研究TPK1，cAMP依赖性 蛋白激酶催化亚基的酿酒酵母。 现有的TPK1晶体，其分辨率为2.2 A， 用重原子化合物衍生，解决晶相 问题，并产生其三维结构。 该结构将 揭示了先前证明涉及的关键残基的位置 在底物识别和催化中的作用，并揭示了 蛋白激酶结构域。 这项工作将通过预测 底物不同于TPK1其它蛋白激酶的结构 使用比较建模方法的特异性。 产生的预测 蛋白激酶底物的选择性将在体外进行测试， 诱变 嵌合激酶将被合成和表征， 将计算机模拟预测的结构特征与酶促反应相关联， 活动 对蛋白激酶结构的全面了解将加快 可能用于治疗的特异性抑制剂的开发 肿瘤性疾病