A Strategy for the Synthesis of N-Linked Glycoproteins

蛋白质的糖基化在许多过程中发挥着重要作用，从蛋白质折叠到 对癌症等疾病的认识。尽管它很重要，但人们对它的影响知之甚少 在分子水平上的蛋白质糖基化。这是由于与隔离相关的困难。 均一的糖蛋白。下面的提案将描述一种新的合成方法 均一的N-连接的糖蛋白。这种方法的第一部分涉及进化突变的trna。 用氨基酸N-乙酰氨基葡萄糖-天冬酰胺(GlcNAc-ASN)充电突变的tRNA的合成酶 以便将其整合到体内的蛋白质中。然后，该提案将描述 具有增强的转糖基化特性的氨基葡萄糖苷酶，用于转移络合物 碳水化合物转变为含有GlcNAc-ASN部分的蛋白质。最后，介绍了自动化一锅锅的应用 低聚糖的合成为突变株构建复合碳水化合物底物 将描述内切葡萄糖胺酶。如果成功，上述方法的组合将 为高效构建均一的N-连接糖蛋白文库提供了新的策略。

Glycosylation of proteins plays a major role in number of processes ranging from protein folding and recognition to diseases such as cancer. Despite its importance there is very little known about the effects of protein glycosylation at the molecular level. This is due to the difficulties associated with isolating homogenous glycoproteins. The following proposal will describe a new approach to the synthesis of homogenous N-linked glycoproteins. The first part of this approach involves evolving a mutant tRNA synthetase to charge a mutant tRNA with the amino acid N-acetyiglucosamine-asparagine (GlcNAc-Asn) in order to incorporate it into proteins in vivo. The proposal will then describe the evolution of a glucosaminidase with enhanced transglycosylation properties, to be used to transfer complex carbohydrates to proteins containing the GlcNAc-Asn moiety. Finally, application of the automated one-pot oligosaccharide synthesis to the construction of a complex carbohydrate substrate for the mutant endoglucosaminidase will be described. If successful the combination of the methods outlined above will provide a new strategy for the efficient construction of libraries of homogenous N-linked glycoproteins.