Fundamental biochemistry of N-glycosylationand the dolichol cycle

将糖添加到蛋白质中是健康人类的一个基本且受到严格监管的过程。N-糖基化是将糖部分加到蛋白质中天冬酰胺残基上的过程，在蛋白质的稳定性、定位和功能方面具有重要的功能，N-糖基化的许多步骤存在缺陷，导致先天性糖基化障碍。用于N-糖基化和其他细胞过程的糖分子被对接到人类细胞内内质网的膜上，这种分子位于一种名为多利胆碱的脂类分子上。许多参与N-糖基化的早期阶段以及多醇载体的合成和回收的酶在文献中仍然没有被描述，大多数工作集中在酵母同系物上。在许多情况下，这是由于难以获得酶或底物。该项目旨在通过制造底物衍生物并将其用于关键酶的生化、生物物理和结构分析来解决这一问题。通过单独观察每个催化剂的机制，以及在复合体中共同作用，我们将评估酶之间的动力学，并建立一个关于早期N-糖基化和多醇循环事件的生化模型。

The addition of sugars to proteins is an essential and highly regulated process in healthy humans. N-glycosylation describes the process in which sugar moieties are added to asparagine residues in a protein, and has important functions in protein stability, localisation, and function, with defects in many of the steps in N-glycosylation leading to congenital disorders of glycosylation. Sugar molecules destined to be used in N-glycosylation as well as in other cellular processes, are docked to the membrane of the endoplasmic reticulum inside human cells, on a lipid molecule called dolichol. Many of the enzymes involved in the early stages of N-glycosylation and in the synthesis and recycling of dolichol carrier remain uncharacterised in the literature, with most work focussing on yeast homologs. In many cases this is due to the difficulty of accessing either the enzyme or substrates. This project aims to address this by making substrate derivatives and using them for biochemical, biophysical and structural analysis of key enzymes. By looking at the mechanisms of each catalyst alone and also together in complexes, we will assess the dynamics between the enzymes and build a biochemical model of events in early N-glycosylation and the dolichol cycle.