Dolichol oligosaccharides

The Bause model also fails to adequately predict the behavior of these substitutions. Their model would seem to predict enhanced turnover from the thioamide, given the increased acidity of the thioamide protons (22). This would also predict that the peptide 8 could be a viable substrate for glycosylation since it could readily transfer a proton to the hydroxy amino acid and form a potent nucleophile. While this peptide binds to the enzyme, no glycosylation or increased hydrolysis of dolichol-oligosaccharide is observed (22). 

Coenzymes serve as recyclable shuttles—or group transfer reagents—that transport many substrates from their point of generation to their point of utilization. Association with the coenzyme also stabilizes substrates such as hydrogen atoms or hydride ions that are unstable in the aqueous environment of the cell. Other chemical moieties transported by coenzymes include methyl groups (folates), acyl groups (coenzyme A), and oligosaccharides (dolichol). 

Dolichol-P-P-oligosaccharide is not involved, nor are gly-cosidases and the reactions are not inhibited by tuni-camycin. 

The Biosynthesis of N-Linked Glycoproteins Involves Dolichol-P-P-Oligosaccharide... 

Before it participates in the biosynthesis of Dol-P-P-oligosaccharide, dolichol must first be phosphorylated to form dolichol phosphate (Dol-P) in a reaction catalyzed by dolickol kinase and using ATP as the phosphate donor. 

Figure 47-7. Pathway of biosynthesis of dolichol-P-P-oligosaccharide. The specific linkages formed are indicated in Figure 47-8. Note that the first five internal mannose residues are donated by GDP-mannose, whereas the more external mannose residues and the glucose residues are donated by dolichol-P-mannose and dolichol-P-glucose. (UDP, uridine diphosphate Dol, dolichol P, phosphate UMP, uridine monophosphate GDP, guanosine diphosphate M, mannose G, glucose.)...

Figure47-8. Structure of dolichol-P-P-oligosaccharide. (Reproduced, with permission, from Lennarz WJ The Biochemistry of Glycoproteins and Proteoglycans. Pienum Press, 1980.)...

The oligosaccharide GICjMangCGIcNAcjj is transferred from dolichol-P-P-oligosaccharide in a reaction catalyzed by oligosaccharide protein transferase, which is inhibited by tunicamycin. 

Tunicamycin Inhibits GIcNAc-P transferase, the enzyme catalyzing addition of GIcNAc to dolichol-P, the first step in the biosynthesis of oligosaccharide-P-P-dolichol... 

Glycosyladon of the p62 afad El proteins is not needed for correct assembly in the ER, since in tunicamycin-treated cells, the assembly of these nonglycosylated proteins proceeds normally (Garoff and Schwarz, 1978). The drug tunicamycin blocks the assembly of the dolichol-linked oligosaccharide intermediate so that no transfer of oligosaccharides to nascent protein is possible (Tkasz and Lampen, 1975). 

In transit through the ER and Golgi, the proteins acquire oligosaccharide side chains attached commonly at serine or threonine residues (O-linked) or at asparagine residues (N-linked). N-linked glycosylation requires participation of a special lipid called dolichol phosphate. 

In the first Section, the dolichol pathway of protein glycosylation is introduced, and the reader is made familiar with the various reactions in the formation of the lipid and carbohydrate moieties of lipid-linked saccharides. Three different classes of compound are known so far (a) isoprenoid alcohol esters of monosaccharide monophosphates, such as D-mannosyl and D-glucosyl (dolichol phosphate), (b) such isoprenoid alcohol esters of saccharide diphosphates as dolichol diphosphate linked to 2-acetamido-2-deoxy-D-glucose and to oligosaccharides, and (c) retinol (D-mannosyl phosphate). The dolichol-linked sugars occur in all eukaryotes. 

In Section II (on the biosynthesis of oligosaccharides), emphasis is placed on the biosynthesis of the dolichol-linked sugars, because the inhibitors mainly interfere with steps in the dolichol cycle of protein glycosylation. Modification of the oligosaccharide side-chain, once it has been transferred from the carrier lipid to the protein, is not discussed in any great detail. 

How, exactly, retinol affects protein glycosylation is not yet known, but the formation of retinol phosphate, the D-mannosylation of retinol phosphate by GDP-Man, and the transfer of Man from D-mannosyl (retinol phosphate) to pronase-sensitive glycoconjugates are well established.5857,59 On the other hand, the formation and role of other gly-cosyl esters of retinol phosphate are still controversial.2,80 It is very probable that the D-mannosylation of retinol phosphate, and also the transfer of Man from this lipid derivative to protein, are catalyzed by enzymes different from those involved in the dolichol pathway,81-83 and this finding points to the different physiological roles of Man-P-Dol and D-mannosyl (retinol phosphate). Thus, whereas Man-P-Dol can serve to D-mannosylate dolichol diphosphate-linked oligosaccharides (see Section III), such a role has not been found for D-mannosyl (retinol phosphate).81,83... 

The biological function of this dolichol-linked oligosaccharide was revealed in pulse-chase experiments which showed that the intact oligosaccharide was transferred, as such, to protein. Subsequent studies, to be discussed, confirmed that the formation of the oligosaccharide linked to the Asn residues of glycoproteins is probably initiated by transfer of the oligosaccharide from the dolichol diphosphate intermediate to the asparagine residue of a nascent polypeptide (see Refs. 35, 49, 50, and 67 for reviews). 

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