Structural Determinants of Glycogen Synthase Regulation

描述（由申请人提供）：本提案的总体目标是了解负责葡萄糖储存聚合物糖原的合成和延伸的酶的结构和功能特性。糖原是葡萄糖的重要储存储备，从酵母到人类，糖原合成的启动和延长机制是保守的。我们最近对酵母糖原合酶的结构研究为了解所有真核酶的功能特性提供了重要的新见解。本提案的长期目标是在初步工作的基础上，描绘糖原合成酶催化反应的结构背景，并揭示糖原结合与糖原合成酶的葡萄糖-6-磷酸活化之间的结构相互作用。作为这项工作的一部分，我们将初始化筛选人类糖原合成酶的小分子调节剂，这些调节剂可以在2型糖尿病中发现的条件下操纵酶的活性状态，促进葡萄糖的处理，或抑制糖原合成酶的作用，作为治疗糖原储存障碍的潜在方法，如Lafora和Pompe病。糖原合成酶及其底物与葡萄糖-6-磷酸结合的复合物或激活剂结合位点的新型激动剂/拮抗剂将提供重要的结构/活性关系，这将指导我们的研究进一步发展。作为Aim 2中筛选工作的一部分，我们将生产代表均质磷酸化状态的新型酶形式，并将针对该目标中确定的糖原合成酶的新型调节剂进行测试。此外，我们将在存在和不存在底物、底物类似物或葡萄糖-6-磷酸的情况下对这些新型调节剂进行结构测定，这将与我们在目标1中的工作有关。目的3将确定催化的分子机制和糖基转移的特异性。该目的的目标之一是了解糖原合成酶产生的催化错误是否最终导致糖原加工畸变。该目标将利用目标1中开发的结构信息，并将使用酵母Gsy2p和人糖原合成酶的野生型和突变型，并将为我们的调节剂发现和开发提供重要的结构/活性信息，作为目标2的一部分。

DESCRIPTION (provided by applicant): The overall goal of this proposal is to understand the structural and functional properties of the enzymes responsible for the synthesis and elongation of the glucose storage polymer, glycogen. Glycogen is an important storage reserve of glucose and the mechanism by which the synthesis of glycogen is initiated and elongated is conserved from yeast to humans. Our recent structural work on yeast glycogen synthase has provided important new insight into the functional properties of all eukaryotic enzymes. The long-range goal of this proposal is to build off this initial work to delineate the structural context for the reaction catalyzed by glycogen synthase and uncover the structural interplay between glycogen association and glucose-6- phosphate activation of glycogen synthase. As part of this effort, we will initialize a screen for small molecule modulators of human glycogen synthase that can manipulate the activity state of the enzyme and promote glucose disposal under conditions found in Type 2 Diabetes or inhibit the action of glycogen synthase as a potential approach for treating glycogen storage disorders, such as Lafora and Pompe's Disease. Structure determination will be an integral part or our efforts as complexes between glycogen synthase and its substrates in combination with glucose-6-phosphate or novel agonists/antagonists of the activator binding site will provide important structure/activity relationships that will guide further development of our research. As part of our screening efforts in Aim 2 we will produce novel enzymatic forms representing homogenous phosphorylation states and these will be tested against the novel modulators of glycogen synthase identified in this aim. In addition, we will pursue structure determination of these novel modulators in the presence and absence of substrates, substrate analogs or glucose-6-phosphate which will link to our work in aim 1. Aim 3 will determine the molecular mechanism of catalysis and the specificity of glycosyltransfer. One of the goals of this aim is to understand whether catalytic mistakes produced by glycogen synthase are ultimately responsible for aberrations in glycogen processing. This aim will draw upon the structural information developed in aim 1 and will employ wild-type and mutant forms of both yeast Gsy2p and human glycogen synthase and will feed important structure/activity information into our modulator discovery and development as part of aim 2.