ROLE OF N-LINKED GLYCANS AND PROTEIN FOLDING

蛋白质折叠是一个基本的过程，但仍然知之甚少。 活细胞。本申请中提出的实验是设计的 要评估折叠领域中一个新概念的分子基础： N-连接的糖链作为伴侣结合的附件的直接作用 在内质网中的糖蛋白折叠和组装过程中。初步结果 表明Calnexin是一种普遍存在的内质网伴侣蛋白，与其 凝集素样亲和部分葡萄糖修剪的N-连接的配体 低聚糖(Glc1Man9-7GlcNAc2)。对于许多糖蛋白来说，这 结合(受葡萄糖苷酶和管腔UDP调节- GLC：糖蛋白葡萄糖转移酶)可能决定 折叠和出口到高尔基综合体的效率。 我们的总体目标是分析钙粘蛋白及其可溶性蛋白的作用。 同源钙网织蛋白在构象成熟度和质量中的作用 控制新合成的糖蛋白。我们将分析 糖蛋白与钙粘蛋白和钙粘连蛋白结合的特异性和结构基础 钙网织蛋白，内质网腔内的一种可溶的同系物。我们还将确定 钙粘蛋白和钙网织蛋白结合的功能意义 在糖蛋白生物发生方面。最后，我们将研究UDP的作用- 葡萄糖：糖蛋白葡萄糖转移酶及其作用机制 在内质网中支持的重新和脱糖循环中。这个 实验将利用生化、细胞生物学、遗传学和 结构性方法。 这一结果将有助于阐明各种不同的 糖蛋白获得继承性转运能力的三维结构 活细胞内质网的结构。结果将直接影响到 内质网储存/质量控制与多种疾病的相关性 病因包括囊性纤维化和α1抗胰蛋白酶缺乏症。这个 需要共翻译糖基化和核心多糖的修剪可能， 此外，在糖基化和糖基化之间的联系中找到它们的解释 折叠、低聚物组装、降解和质量控制。

Protein folding is a fundamental but still poorly understood process in living cells. The experiments proposed in this application are designed to evaluate the molecular basis of a novel concept in the folding field: the direct role of N-linked glycans as appendages for chaperone-binding during glycoprotein folding and assembly in the ER. Preliminary results indicate that calnexin, a ubiquitous ER chaperone, associates with its ligands by lectin-like affinity to partially glucose-trimmed N-linked oligosaccharides(Glc1Man9-7GlcNAc2). For many glycoproteins, this association (modulated by glucosidases and the lumenal UDP- Glc:glycoprotein glucosyltransferase) is likely to determine the rate of folding and the efficiency of export to the Golgi complex. Our overall goal is to analyze the role of calnexin and its soluble homologue calreticulin in the conformational maturation and quality control of newly synthesized glycoproteins. We will analyze the specificity and structural basis for glycoprotein binding to calnexin and calreticulin, a soluble homologue in the ER lumen. We will also determine the functional significance of the calnexin and calreticulin association in glycoprotein biogenesis. Finally, we will investigate the role of UDP- glucose: glycoprotein glucosyl transferase, and the mechanisms involved in the re-and deglucosylation cycle that it supports in the ER. The experiments will utilize biochemical, cell biological, genetic and structural approaches. The results will throw light on the mechanisms by which a variety of glycoproteins acquire heir transport-competent three-dimensional structure in the ER of living cell. The results will have direct relevance to a variety of diseases with ER storage/quality control etiology including cystic fibrosis and alpha1-antitrypsin deficiency. The need for cotranslational glycosylation and trimming of core glycans may, moreover, find their explanation in the link between glycosylation and folding, oligomeric assembly, degradation and quality control.