Cellular protein maturation and degradation

这项提案的总体目标是确定N-连接的多糖在 穿越分泌途径的蛋白质的成熟和质量控制。这个 大多数通过分泌途径传输的蛋白质都会收到多个N-连接的 糖基化作用。我们和其他人已经证明，这些碳水化合物被用作 成熟度和质量控制信号，以传达关于 成熟的新生链条，并有助于决定它们的命运。有关的信息 蛋白质的构象在N-连接的葡聚糖的组合物中由一种 内质网(ER)中的一系列糖苷酶和转移酶。 这些编码蛋白能够根据糖链调节糖链的组成。 蛋白质的结构。UDP-葡萄糖：糖蛋白葡萄糖转移酶(GT1)是 目前已知的唯一一种可以改变糖链结构的蛋白质 关于修饰蛋白质的性质，人们对其在活细胞中的活性知之甚少。这个 然后，糖的组成被识别的碳水化合物结合蛋白破译 不同的多糖结构，作为分子伴侣或分类受体来协助 在成熟和质量控制过程中。碳水化合物的葡萄糖臂 支持凝集素伴侣钙粘蛋白和钙网织蛋白的结合，从而指导其协同- 和平移后折叠过程。相比之下，甘露糖枝似乎 在引导蛋白质从内质网中退出方面起着重要作用。天然蛋白质是 送到高尔基体，而异常蛋白质则由胞质进行分类销毁 蛋白酶体通过内质网相关蛋白降解(ERAD)途径。因此， 甘露糖苷酶似乎参与了质量控制代码的生成，即 由碳水化合物结合的分选受体识别，控制着 来自急诊室的蛋白质。这种修饰和识别多聚糖的内质网 基于修饰蛋白的构象，它在辅助 分泌物中糖蛋白的成熟过程及导向质控运输 路径。我们的具体目标是：(1)了解多糖的作用和载体性质 细胞折叠反应在丙皂苷的有效成熟中的作用 含结构域蛋白；(2)阐明GT1在成熟期和成熟期的功能。 GT1底物丙皂苷的质量控制；以及(3)确定丙皂苷的作用 甘露糖苷酶样蛋白EDEM1在非天然蛋白分选和递送中的作用 内质网相关蛋白降解途径。

The overall goal of this proposal is to determine the role of N-linked glycans in the maturation and quality control of proteins that traverse the secretory pathway. The majority of proteins that travel through the secretory pathway receive multiple N-linked glycosylations. We and others have shown that these carbohydrates are used as maturation and quality control signals to convey information about the fitness of maturing nascent chains and help to determine their fate. Information about the conformation of a protein is encoded within the composition of the N-linked glycan by a series of glycosidases and transferases that reside in the endoplasmic reticulum (ER). These encoder proteins are able to modulate the glycan composition based on the structure of the protein. The UDP-glucose: glycoprotein glucosyltransferase (GT1) is currently the only known protein that can modify the glycan structure dependent upon the properties of the modified protein, yet little is known about its activity in live cells. The glycan composition is then deciphered by carbohydrate-binding proteins that recognize distinct glycan structures and act as molecular chaperones or sorting receptors to assist in maturation and quality control processes. The glucose arm of the carbohydrate supports binding by the lectin chaperones calnexin and calreticulin, which direct its co- and post-translational folding processes. In contrast, the mannose branches appear to serve important roles for directing the exit of proteins from the ER. Native proteins are sent to the Golgi, whereas aberrant proteins are sorted for destruction by the cytosolic proteasome through the ER-associated protein degradation (ERAD) pathway. Therefore, mannosidases appear to be involved in generating the quality control code that is recognized by carbohydrate-binding sorting receptors that control the trafficking of proteins from the ER. This ER network of enzymes that modify and recognize glycans based on the conformation of the modified protein plays a crucial role in assisting the maturation process and directing quality control traffic of glycoproteins in the secretory pathway. Our specific aims: (1) to understand the role of glycans and the vectorial nature of the cellular folding reaction in the efficient maturation of prosaposin, a sequential domain containing protein; (2) to elucidate the function of GT1 in the maturation and quality control of prosaposin, an obligate GT1 substrate; and (3) to determine the role of the mannosidase-like protein, EDEM1, in the sorting and delivery of non-native proteins to the ER-associated protein degradation pathway.