REGULATION OF PROTEIN N GLYCOSYLATION IN THE CNS

描述：蛋白质N-糖基化是蛋白质组装的关键阶段。 膜糖蛋白介导的重要神经生物学功能， CNS。 N-连接的寡糖链最初合成为 脂质连接前体寡糖，Glc3Man9GlcNAc2-P-P-多萜醇 （寡聚-P-P-Dol）通过多阶段组装过程， Man5GlcNAc 2-P-P-Dol、甘露糖基磷酸肌醇（Man-P-Dol）的形成 和葡糖基磷酸多萜醇（Glc-P-Dol）的细胞质面上的细胞质。 内质网（ER），糖核苷酸作为糖基 捐助者。 生物合成完成后，极性磷酸糖头 三种脂质中间体的组横向扩散到管腔 单层 尽管糖基生物合成的许多细节 载体脂质，磷酸多里酯（Dol-P）和寡聚-P-P-Dol已被 尽管已经阐明，但在理解 参与Dol-P生物合成的酶的数量和调节 从头开始和三个脂质中间体的跨双层运动。 以来 已经确定，长链水平的增加 顺式异戊二烯基转移酶（cis-lPTase）系统，催化链 在Dol-P生物合成的延伸阶段，是一个关键的调控事件， 在发育中的脑细胞中诱导N-糖基化装置， 申请提出了广泛的新的酶学和分子研究， 顺式-lPTase系统。 该申请的另一个主要方面涉及 关于膜蛋白的基本问题假定介导的 Man-P-Dol、Glc-P-Dol和Man5GlcNAc 2-P-P-Dol在细胞膜中的跨双层运动 儿 该项目的这一部分扩展了Man-P-香茅醇 （Man-P-Cit）是Man-P-Dol的水溶性类似物， 通过蛋白质介导的系统密封ER囊泡。 一个主要目标是 确定介导Man-P-Cit转运的蛋白质与 Man-P-Dol "翻转酶"，并探讨其他相关的使用 水溶性类似物用于类似研究。 拟议的实验将 提供有关结构和监管的重要新信息， 顺式-lPTase系统在大脑中的作用，并有助于了解 极性脂质在ER中横向扩散的机制， 膜生物学中最重要的问题。

DESCRIPTION: Protein N-glycosylation is a critical stage in the assembly of membrane glycoproteins mediating important neurobiological functions in the CNS. N-linked oligosaccharide chains are initially synthesized as the lipid-linked precursor oligosaccharide, Glc3Man9GlcNAc2-P-P-dolichol (Oligo-P-P-Dol) by a multi-stage assembly process that begins with the formation of Man5GlcNAc2- P-P-Dol, mannosylphosphoryidolichol (Man-P-Dol) and glucosylphosphoryldolichol (Glc-P-Dol) on the cytoplasmic face of the endoplasmic reticulum (ER) with sugar nucleotides serving as the glycosyl donors. Biosynthesis is completed after the polar phosphosaccharide head groups of the three lipid intermediates diffuse transversely to the lumenal monolayer. Although many details of the biosynthesis of the glycosyl carrier lipid, dolichyl phosphate (Dol-P), and Oligo-P-P-Dol have been elucidated, there are still many crucial gaps in the understanding of the number and regulation of the enzymes involved in the biosynthesis of Dol-P de novo and the transbilayer movement of three lipid intermediates. Since it has been established that increased levels of the long chain cis-isoprenyltransferase (cis-lPTase) system, catalyzing the chain elongation stage in Dol-P biosynthesis, is a key regulatory event in the induction of the N-glycosylation apparatus in developing brain cells, this application proposes extensive new enzymological and molecular studies on the cis-lPTase system. Another major aspect of the application addresses basic questions about the membrane proteins postulated to mediate the transbilayer movement of Man-P-Dol, Glc-P-Dol and Man5GlcNAc2-P-P-Dol in the ER. This part of the project extends the finding that Man-P-citronellol (Man-P-Cit), a water-soluble analogue of Man-P-Dol, is transported into sealed ER vesicles by a protein-mediated system. A key objective is to establish that the protein(s) mediating Man-P-Cit transport is related to Man-P-Dol "flippase", and to explore the use of other pertinent water-soluble analogues for similar studies. The proposed experiments will provide important new information about the structure and regulation of the cis-lPTase system in brain, and contribute to the understanding of the mechanisms by which polar lipids diffuse transversely in the ER, a fundamentally important problem in membrane biology.