Chemical Synthesis And Properties Study Of Glycosylation Polypeptide And Protein

Protein is one of the main constituents of the living body,the complete primary sequence of polypeptide chain and accurate three-dimensional structure of protein is the basis of its normal functions.The change of protein structure and post-translational modification usually connects with some disease.In addition to the factors of protein sequence itself,post-translational modifications are the most important factors affecting protein structure.The glycosylation is one of the most common type.There are several possible glycosylation sites and the glycans have complex structures.In order to study the influence of glycosylation on protein structure more accurately,we have to get a specific site modified form homogeneous protein.It is difficult to get this kind of protein from nature.There are also some difficulties for synthetic method to obtain complex modified protein,however the syntheticed proteins have a definite sequence and modified.Synthetic method can provide accurate research model for researchers.How to complete the synthesis of difficult sequences,the development of new synthetic methods and strategies are problems needed to be solved.In this paper,we choose disease-related glycopeptides and glycoproteins as the research object and use solid phase peptide synthesis,protein expression and natural chemical ligation（NCL）and other methods to investigate the synthesis method and protein synthesis strategy for difficult glycosylated polypeptides and protein.In this paper,we utilize solid phase peptide synthesis,protein expression and native chemical ligation methods to finish the synthesis of diseased related glycopeptide and glycoproteins.We found that the glycosylation sites influenced the immune activity of the vaccines,but the TLR2 ligand chiral has little impact.Firstly,we synthesized glycopeptide based on repeat variable sequence in MUC1protein,and identified immunogenicity and specificity of the glycopeptide.Then we covalently conjugate the glycopeptide with Toll like receptor ligand Pam₃CysSerLys₄to build the vaccine and evaluate its immunity.In addition,we explored the synthesis methods and strategies for glycosylated murine Prion protein.Prion is easy to form precipitation and hydrophobic aggregates,It makes a lot problems for protein synthesis and purification.In order to complete the synthesis of Prion.We take the murine Prion sequence 90-230 as the research objectives.We combine the biological expression and chemical synthesis method.The N-terminal of this protein sequence 90-177 was obtained by prokaryotic expression.We introduce a cysteine in the end of the natural sequence for the syntheis of protein thioester.The glycopeptide of the C-terminal was obtained by chemical synthesis.We try to use the isopeptide building blocks and main chain protecting groups to complete the synthesis of this part of the difficult sequence.At the same time we also synthesized polypeptide fragments with the glycosylation modification.Finally,we use the natural chemical ligation method to build the murine Prion sequence 90-230 protein.