STRUCTURE-FUNCTION STUDIES OF 70 KD HEAT SHOCK PROTEINS

我们提出了一系列的结构和力学研究 70千道尔顿(KDa)热休克样蛋白成员 一家人。根据其中几种蛋白质的序列数据， 蛋白质家族。基于其中几个的序列数据 蛋白质，以及对牛笼状蛋白功能的描述 去涂层ATPase(HSC70)，我们提出了一个工作模式：全部70 KDA热休克蛋白从一个共同的祖先进化而来， 维持高度保守的ATPase结构域并发展 专门的底物识别域。他们都应该 相似的三级结构，而且都可能催化 使用其进行某种类型的大分子重排 根本机制。 我们将开始对这一蛋白的代表进行研究 生物化学功能为人所熟知的家庭， 牛HSC70。我们将解开x射线晶体结构。 其44 kDa的ATPase核心，并将尝试结晶和 解开完整的HSC70的结构。我们正在克隆，并将 HSC70mRNA的序列；我们将尝试 构建该蛋白的重组高产表达载体。我们会 对HSC70进行蛋白质修饰(突变)实验 为了解决衬底识别的机理问题和 构象变化。 第二位代表，我们将对其进行类似的研究 Tandem是牛葡萄糖调节蛋白。

We propose a series of structural and mechanistic studies on members of the 70 kilodalton (kDa) heat-shock-like protein family. Based on sequence data for several of these proteins, protein family. Based on sequence data for several of these proteins, plus delineation of the function of the bovine clathrin uncoating ATPase (HSC70), we propose a working model: all 70 kDa heat shock proteins evolved from a common ancestor, maintaining a highly conserved ATPase domain and developing specialized substrate recognition domains. They all should have similar tertiary structures, and all probably catalyze macromolecular rearrangements of some type using the same fundamental mechanism. We will initiate our studies on the representative of this protein family for which a biochemical function is well-understood, bovine HSC70. We will solve the x-ray crystallographic structure of its 44 kDa ATPase core, and will attempt to crystallize and solve the structure of the intact HSC70. We are cloning and will sequence cDNA for the HSC70 mRNA; we will attempt to construct a recombinant overproducer of the protein. We will pursue protein modification (mutagenesis) experiments on HSC70 to address mechanistic questions of substrate recognition and conformational change. A second representative on which we will pursue similar studies in tandem is the bovine glucose regulated protein.