Williams BCH 444 2014 Lectures 3-6

G cell surface or secreted sialic acid n

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Unformatted text preview: saccharides found on mature glycoproteins, e.g., cell surface or secreted Sialic acid N-acetylgalactosamine (GalNAc) biantennary high mannose triantennary tetraantennary complex forms What determines the types of structures found at a particular glycosylation site in a glycoprotein? 1)  Competition between glycosyltransferases for a common substrate. Often the product of one enzyme inhibits the action of others - commits to various pathways 2)  Inaccessibility of an oligosaccharide to glycosyltransferases due to protein folding. Primary reason why high mannose precursor structures are found on mature glycoproteins 3) Biosynthetic rates - rate at which glycoprotein passes through Golgi. Fast rate – less opportunity for transferases to act 4) Substrate concentrations, e.g., one type of nucleotide sugar donor may be at higher concentration than others Microheterogeneity - a given site on different molecules of a particular glycoprotein can have different N-glycans that differ, for example, in branching Control of branching by GlcNAc transferases 42 So many structures, so much energy used - what s the point?     Oligomerization through disulfide cross-linking (molecular weights > 1 MegaDa) 43 Glycoprotein Functions II Mol. Biol Cell 5th Ed Major types of animal lectins. The various carbohydrate-recognition domains (CRDs) shown: (CL) C-type lectin CRD; (GL) galectin CRD; (MP) P-type lectin CRD; (IL) I44 type lectin CRD. Other domains are (EG) EGF-like domain; (IG2) immunoglobulin C2-set domain; (TM) transmembrane region; and (C3) complement regulatory repeat. Glycoprotein Functions III - Role of lectins in the secretory pathway Targeting misfolded proteins for degradation by the ER lectins EDEM and OS9 (specificity Man6-8GlcNAc2) Protein folding assisted by the ER lectin-chaperones calnexin and calreticulin (specificity: Glc1Man5-9GlcNAc2) Targeting a subset of glycoproteins for export from the ER by lectins ERGIC-53 and VIP36 (specificity uncertain - high mannose oligosaccharides) Targeting lysosomal enzymes from trans-Golgi to lysosomes by mannose-6-P receptor (lectin) 45 Glycoprotein Functions III - Role of lectins in the secretory pathway (lysosomal targeting) (GlcNAc phosphodiester glycosidase located mainly in trans-Golgi) Lysosomal hydrolase arrives in cis-Golgi with Man8GlcNAc2 oligosaccharides How is it distinguished from secretory glycoproteins with the same oligosaccharide? Once Man-6-P signal is uncovered, it is recognized by the P-type lectin Man-6-phosphate receptor (MPR) I-cell disease: Inherited deficiency in GlcNAc phosphotransferase. Patients have build up of undigested material in lysosomes and their lysosomal hydrolases are secreted rather than targeted to lysosomes. 46 Glycoprotein Functions IV - Role of lectins as cell adhesion molecules Tethering of circulating leukocytes to activated endothelium via interactions between selectins and their ligands In inflamed vessels, selectins and integrin ligands are expressed on endothelial surfaces. This leads to tethering, rolling, and arrest of circulating leukocytes and their eventual extravasation from the circulation to the surrounding tissue. Selectins are specific for sialyl Lewis X structure on O-linked oligosaccharides (up to 70 on PGSL-1) 47 Glycoprotein Functions IV - Role of lectins in defense against pathogens 48 Glycoprotein Functions IV - Role of lectins in attack by pathogens HA NA NA Sugar-dependent steps in the influenza relication cycle: (STEP 1) A virion attaches to sialic acid on the host cell membrane via hemagglutinin (HA) and enters the cytoplasm by receptor-mediated endocytosis, thereby forming an endosome. (STEP 7) The newly synthesized virions bud from the infected cell. Viral neuraminidase (NA) destroys the sialic acid moiety of cellular receptors, thereby releasing the progeny virions. 49 Sugar-based antivirals - Tamiflu neuraminidase inhibitor Sialic acid Tamiflu (oseltamivir) Structure of Tamiflu bound to the active site of viral neuraminidase (monomer and tetramer) Structure-based drug design by Gilead Sciences and marketed by Hoffmann-La Roche 50 Second Quiz extracellular 8765432 1 cytosol –OOC NH3+ 51...
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