Effects of glycosylation on protein structure, function and dynamics

近年来，蛋白质结构生物学研究取得了显著进展。然而，具有翻译后修饰（如糖基化、磷酸化和甲基化等）的人蛋白或复杂蛋白在蛋白质数据库（PDB）中很少发现，因为这些修饰不能由大肠杆菌等原核蛋白表达宿主充分完成。在翻译后修饰中，蛋白质糖基化是自然界中发现的最丰富的蛋白质修饰，它比其他所有翻译后修饰加起来给蛋白质带来更多的多样性。蛋白质的糖基化改变了它们的结构、热力学和动力学，从而调节了蛋白质的功能，影响它们的定位、运输、溶解度、抗原性、生物活性、半衰期和细胞间相互作用。因此，聚糖结构和糖基化模式在发育过程和某些疾病中是高度动态和变化的，这并不奇怪。到目前为止，人们对蛋白质糖基化对功能的影响知之甚少。更重要的是，关于这种翻译后修饰的结构信息很少。事实上，从结构生物学的角度来看，糖基化是不希望发生的，由于低聚糖链的大小、灵活性和异质性，糖基化通常被认为是解决蛋白质结构的额外障碍。我们课题组（P6 Schwalbe）希望通过核磁共振光谱研究糖基化对糖蛋白结构、功能和动力学的影响。为此，我们将在第一个资助期内开发特定的核磁共振工具（表达，同位素标记，核磁共振方法）来进行糖基化蛋白的研究。在研究单元中，我们将重点关注三种高度保守的糖基化途径（n -糖基化，c -甘露糖基化和o -甘露糖基化），它们基于脂质醇，并竞争甘露糖基供体底物和受体蛋白。在这个研究单元的组装过程中，我们已经开始与（P1 Bakker）合作，通过核磁共振对色氨酸c -甘露糖基化的影响进行结构研究。

Recent years have seen remarkable progress in the field of structural biology of proteins. However, human or complex proteins with posttranslational modifications (like glycosylation, phosphorylation and methylation etc.) are rarely found in the Protein Data Bank (PDB), since these modifications cannot be performed sufficiently by prokaryotic protein expression hosts like Escherichia coli. Among the posttranslational modifications, protein glycosylation is the most abundant protein modification found in nature, introducing more diversity into the protein than all the other posttranslational modifications combined. Glycosylation of proteins changes their structure, thermodynamics and kinetics, which modulate the function of the proteins and influence their localization, trafficking, solubility, antigenicity, biological activity, half-life and cell-cell interactions. Therefore, it is not surprising that glycan structures and glycosylation patterns are highly dynamic and change during development and in certain diseases. So far, the effects of protein glycosylation on function are poorly understood. Even more, structural information regarding this posttranslational modification is rare. In fact, from a structural biology perspective glycosylation is undesired and is often considered as an additional obstacle to solve a protein structure due to the size, flexibility and heterogeneity of the oligosaccharide chains. In the Research Unit our group (P6 Schwalbe) wants to study the influence of glycosylation on the structure, function and dynamics of glycoproteins by NMR spectroscopy. For this purpose, we will develop specific NMR tools (expression, isotope labeling, NMR methods) to conduct research on glycosylated proteins in the first funding period. In the Research Unit we will focus on three highly conserved glycosylation pathways (N-glycosylation, C-mannosylation and O-mannosylation) which are based on the lipid dolichol and compete for both mannosyl donor substrates and acceptor proteins. In the course of the assembly of this Research Unit we already started to structurally investigate the effect of tryptophan C-mannosylation by NMR in collaboration with (P1 Bakker).