Dolichol phosphates

FIGURE 8.18 Dolichol phosphate is an initiation point for the synthesis of carbohydrate polymers in animals. The analogous alcohol in bacterial systems, undecaprenol, also known as bactoprenol, consists of 11 isoprene units. Undecaprenyl phosphate delivers sugars from the cytoplasm for the synthesis of cell wall components such as peptidoglycans, lipopolysaccharides, and glycoproteins. Polyprenyl compounds also serve as the side chains of vitamin K, the ubiquinones, plastoquinones, and tocopherols (such as vitamin E). 

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 47-6. The structure of dolichol. The phosphate in dolichol phosphate is attached to the primary alcohol group at the left-hand end of the molecule. The group within the brackets is an isoprene unit (n = 17-20 isoprenoid units).

If the synthesis starts from glucose molecules, then the initial step is the transfer of glucose residues from UDP-glucose onto an intermediary acceptor-dolichol phosphate (membrane-bound polyprenol phosphate). Dolichol phosphate assists in the synthesis of an... 

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. 

R7. Richards, J. B., Evans, P. J., and Hemming, F. W., Dolichol phosphates as acceptors of mannose from guanosine diphosphate mannose in liver systems. Biochem. J. 124, 957-959 (1971). 

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. 

The initial demonstration that isopentenyl diphosphate may be used in the biosynthesis of dolichol phosphate in animals and plants... 

Scheme 1.—Pathway of the Biosynthesis oi Dolichol Phosphate and ot Doliehol-1 inked Saccharides. (Reactions requiring more than one enzyme are indicated by dashed arrows. The Scheme is based on work cited in Refs. 1, 3, 8—25, 35, and 50.)...

As shown in Scheme 1, two other pathways lead to the formation of dolichol phosphate. These are (a) phosphorylation of the free poly-prenol, and (b) dephosphorylation of dolichol diphosphate. A dolichol phosphate-cleaving phosphatase has been described as occurring in Tetrahymena pyriformis,15 human lymphocytes,18 calf brain,17 and rat-nerve tissue.18 The protozoal enzyme was a soluble acid phosphatase, although it was clearly different from the bulk phosphatase activity, whereas the mammalian enzymes had optima at neutral pH and were membrane-bound. 

A particulate, enzyme preparation from human lymphocytes contains a phosphatase that converts dolichol diphosphate into dolichol phosphate and phosphate.15 This alkaline phosphatase may play a role in the recycling of dolichol phosphate, if this compound is to act as a coenzyme. However, the specificity of the phosphatase for dolichol diphosphate, 2,3-dehydrodolichol diphosphate, or dolichol diphosphate-linked oligosaccharides27 has not yet been determined. 

The finding that dolichols differ in chain length has, of course, raised the question of the polyprenyl specificity of the various glyco-syltransferases. The idea that different (that is, not miscible) pools of dolichol phosphate exist for different glycosyltransferases has also emerged, and was discussed in a review by Elbein.35 Conflicting evidence has been obtained,36-38 and this point needs further investigation. 

The enzymes catalyzing these reactions are membrane-bound, and are stimulated by exogenous dolichol phosphate. It has been suggested, and now shown43 (at least in the formation of GlcNAc-PP-Dol by enzymes from yeast), that Dol-P stimulates the incorporation of sugar residues into the lipid intermediates because it serves as a substrate. 

From bisubstrate, kinetic analysis with a transferase from hen oviduct that, under the conditions of the assay, formed only GlcNAc-PP-Dol, it followed that both dolichol phosphate and UDP-GlcNAe have to he bound to the enzyme before release of the product occurs.52 However, the fact that only partially purified preparations have thus far been obtained (the preparations may also still be contaminated with substrates and product), together with experimental difficulties in handling both the substrate dolichol phosphate (which, furthermore, is not one compound, see the earlier discussion) and the unstable enzyme (enveloped in micelles of detergent), make difficult a sensible interpretation and comparison of the kinetic parameters detenuined for the different enzvme-preparations. The solubilized enzymes catalyzing reactions 1,2, and 3 have in common their alkaline pH optima and dependence on Mg2+ or Mn2+ ions. The latter fact makes (ethylenedinitrilo)tetraacetic acid (EDTA) a reversible inhibitor of enzyme activity and an important experimental tool. 

CDG-Ij UDP-GlcNAc dolichol phosphate N-acetylglucos-aminephosphotransferase 1 DPAGT1 1 lq23 608093... 

Scheme 2. —Proposed Scheme for the Reactions Involved in the Synthesis of Cellulose in the Green Alga Prototheca zopfii (After Hopp and Coworkers141). [Abbreviations used are Dol = dolichol, Glc = D-glucose, Dol-P = dolichol phosphate, -P-P = diphosphate, and P, = inorganic phosphate.]...

All N-linked oligosaccharides include a common core of five sugars linked to Asn Mana(l,3)Mana(l,6)Man 6-(l,4)GlcNAc/3(l,4)GlcNAc/31,Asn. This reflects a common biosynthetic pathway in which sugars are added to a membrane-associated polyprenol lipid called dolichol phosphate... 

The structure of dolichol phosphate. The hydrocarbon portion of the molecule has a high affinity for membrane structures. The phosphate end forms an activated complex with oligosaccharide intermediates. 

The structure of undecaprenol phosphate. Isoprene phosphates such as undecaprenol phosphate are important carriers and activators in the synthesis of oligosaccharides. Note the similarities to dolichol phosphate. 

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