糖蛋白和糖肽的化学合成教材分析.ppt

Biosynthesis of N-glycoproteins Synthesize of a lipid bound oligosaccharide as the precursors for all the various oligosaccharides expressed in N-glycoproteins Transfer precursor oligosaccharide from the lipid-bound form to an asparagine side chain of a polypeptide chain Trimming and processing of the oligosaccharide Step 1.1Production of GlcNAc-P-P-Dolichol Step 1.2N-Glycan Precursor on the Cytosolic Leaflet of the Endoplasmic Reticulum (ER) Step 1.3 Biosynthesis of the N-Glycan Precursor on Lumenal Leaflet of ER Enzymes Useful in detecting Steps in N-glycan Biosynthesis Carbohydrate Chemistry  Glycobiology Importance of glycoanalysis Pharmaceutical Concerns Regarding Carbohydrates Pharmacokinetics: Influence on receptor binding. Pharmacodynamics: Distribution. Clearance. Define product as “well-characterized”. Lot-to-lot variability. Definition of intellectual property Some Glycoconjugate Disease Associations immunity to infectious diseases, including HIV rheumatoid arthritis (altered composition of IgG and levels of the serum mannose-binding protein) prion diseases congenital disorders of glycosylation (CDGs) (rare, usually resulting in CNS impairment) oral pathologies cystic fibrosis heart pathologies cancer Carbohydrate Analysis Offers Unique Challenges Structural analysis of oligosaccharides is more challenging than that of oligonucleotides or proteins Synthesis is not “template driven”. This is because of the complexities introduced by: Branching Linkages Anomericity Site-specific glycosylation. Microheterogeneity High Sensitivity is essential as the quantities of oligosaccharides are generally low (μg-sub μg) Workflow in glycoanalysis  Effective microisolation of glycoproteins (short LC microcolumns; lectin-based separations; gels and blotting techniques) Sample treatment at microscale (microscale enzymatic and chemical cleavages; sample clean-up; possibly derivatization) Separation of glycan mixtures by CZE, CEC, or capillary LC Sensitive MS meas