PMC

No pentasaccharide-modified dolichol phosphate accumulates in Hfx. volcanii cells lacking AglB nor is detected using aminopropyl resin for liquid chromatography.
A. Tetrasaccharide-modified C55 and C60 dolichol phosphate are detected upon LC-ESI/MS analysis of the total lipid extract from Hfx. volcanii parent strain (upper panel) and ΔaglB cells (lower panel). In neither case is pentasaccharide-modified dolichol phosphate detected. The peaks shown correspond to doubly charged [M–2H]²⁻ ions, with the positions of the different glycan-modified dolichol phosphates indicated.
B. Tetrasaccharide-modified C55 and C60 dolichol phosphate are detected upon LC-ESI/MS analysis of the total lipid extract from Hfx. volcanii parent strain (upper panel) and ΔaglB cells (middle panel) using aminopropyl resin for the liquid chromatography step. In neither case is pentasaccharide-modified dolichol phosphate detected. In contrast, pentasaccharide-modified C55 and C60 dolichol phosphates are detected upon similar analysis of the total lipid extract from Har. marismortui cells (lower panel). The peaks shown correspond to [M–H]⁻ ions, with the positions of the different glycan-modified dolichol phosphates indicated.

Structure of Dolichol and their function during protein N-glycosylation. (A) Structure of Dolichol, where n is dependent on particular cis-prenyltransferase. Dolichol phosphate is an isoprenoid compound (90-100 carbons total) made from dolichol by phosphorylation. Dolichol phosphate performs important functions in synthesis of N-linked glycoproteins, as illustrated in (B). Dolichol phosphate is the structure upon which the complex oligosaccharide is made before transfer to the target protein. The addition of the first moiety, N-acetylglucosamine from UDP-N-acetylglucosamine, can be blocked by the antibiotic, tunicamycin. After assembly of the oligosaccharide is complete, the carbohydrate structure is transferred from dolichol phosphate to an asparagine residue of a target protein having the sequence Asn-x-Ser/Thr, where × is any amino acid. As also shown in (B), Dolichol phosphate can also act as a carrier of sugars to oligosaccharide chain synthesis assembly; such activated sugars include dolichol-P-mannose and dolichol-P-glucose.

De novo biosynthesis of dolichol phosphate in human. Polyisoprenoids are generated by the condensation of isopentenyl pyrophosphate (IPP) and dimethylallyl pyrophosphate (DMAPP), the 5-carbon building blocks of all isoprenoids. The mevalonate pathway is responsible for the synthesis of IPP and DMAPP. The first step dedicated to dolichol biosynthesis is the elongation of farnesyl pyrophosphate by the dehydrodolichyl diphosphate synthase (DHDDS) . This cis-prenyl transferase uses IPP as substrates to produce polyprenol pyrophosphate. It is believe that this polyprenol pyrophosphate is then dephosphorylated into polyprenol before its reduction by SRD5A5. Finally, the dolichol kinase (DOLK) transfers a phosphate from CTP to dolichol

Pentasaccharide-modified dolichol phosphate is detected in a total lipid extract from Har. marismortui but not from Hfx. volcanii. A. LC-ESI/MS analysis of the total lipid extract from Hfx. volcanii (upper panel) and Har. marismortui (lower panel) reveals the presence of tetrasaccharide-modified C55 and C60 dolichol phosphate in both extracts. In contrast, pentasaccharide-modified dolichol phosphate is only seen in the Har. marismortui sample. The peaks shown correspond to [M–2H]²⁻ ions. The positions of the different glycan-modified dolichol phosphates are indicated. B. MS/MS analysis of Har. marismortui pentasaccharide-modified C60 dolichol phosphate. The fragmentation profile is schematically represented in the inset.