Extraction Of Natural Sugar Chain And Synthesis Of Glycopeptides

Glycosylation is one of the most common post-translational modifications of proteins. The heterogeneity of glycan in native glycoproteins and glycopeptides lead to the significant differential charaters in their structure and function. Thus, synthesis of homogeneous glycoproteins and glycopeptides are significant when it comes to study the structure and function relationship of oligosaccharide.In this dissertation, we had explored the method to prepare oligosaccharide substrates from Ribonuclease B(RB), Sialylglycopeptide(SGP) and Ovalbumin(OVA), three natural sources of glycoproteins/glycopeptides. And successfully had we prepared high mannose type and complex type N-glycan oxazoline substrates. After that 10 of GlcNAc containing polypeptides had been selected and synthesized as the acceptors of glycopeptides. We had chemoenzymaticlly synthesized 14 of high mannose type and complex type glycopeptides through the GlcNAc-peptides and glycan oxazolines we had synthesized. We also improved the method of enzymatic synthesis of glycopeptides, established the method to extract high mannose type sugar chain from Ribonuclease B and subsequently prepared the corresponding oxazoline. And had we explored the condition to separate and extract the oligosaccharide from Ovalbumin. Finally the confirmed glycopeptides had been synthesized for the use of Carbohydrate- Proteomic analysis.The methods applied for synthesizing glycopeptides, the study of the ENGase-based chemoenzymatic synthesis of glycopeptide and the application of glycopeptides have been discussed in chapter 1. In chapter 2,the semi-synthesis of glycan oxazoline as the donor substrates have been described. Multiple steps and complicated purification processesmade it difficult to obtain integrated oligosaccharide substrates. Herein, we successfully separated microgram scale oligosaccharide from natural protein resources, and further synthesized the corresponding oxazoline substrates with "one-pot" strategy, in which simple synthesis steps and purification procedures had been developed. Together with the strategy we could obtain transglycosylation substrates quickly and efficiently. In chapter 3, we had synthesized a series of Asn-linked GlcNAc-containing polypeptides, the transglycosylation acceptors. Fmoc solid phase peptide synthesis can make the obtain of polypeptides easily, quickly and efficiently. To obtain homogeneous glycopeptides efficiently, in chapter 4, had we reported the endoglycosidase-catalyzed reaction with which can we attach the synthesized oxazoline substrates to the synthesized polypeptides. And in this chapter, we also explored the method of new oligosaccharide extraction and synthesis of correspondingoxazoline from nature glycoproteins for the obtaining of homogeneous glycopeptides.