Dimethylallyl transferase

Figure 3-14. Possible biosynthetic route towards coumarin. (a) 2-hydroxylase, (b) glucosyl transferase, (c) P-glucosidase, (d) dimethylallyl transferase and/or UV light. The enzymes have not yet been identified.

Either the aglycone resulting from the action of a P-glucosidase or the glucoside, or possibly both, undergo cis-trans isomerization under influence of UV-light or possibly mediated by a dimethylallyl transferase. The last step of the biosynthetic pathway is an intramolecular esterification reaction, which can occur spontaneously, to yield coumarin (3.96). The enzymes that involved in these reactions have not been purified. 

YAMAMOTO, H., SENDA, M., INOUE, K., Flavanone 8-dimethylallyl transferase in Sophora flavescens cell suspension cultures. Phytochemistry, 2000,54,649-655. 

Dhillon, D.S. and Brown, S.A. (1976) Localization, purification and characterization of dimethylallylpyrophosphate umbeUiferone dimethylallyl transferase from Ruta graveolens. Arch. Biochem. Biophys., 177, 74-83. 

Heinstein PF, Lee SI, Floss HG (1971) Isolation of dimethylallylpyrophosphate tryptophan dimethylallyl transferase from the ergot fungus Claviceps spec.). Biochem Biophys Res Commun 44 1244-1251... 

The enzyme responsible for condensation of L-tryptophan and DMAPP is dimethylallylpyrophosphate L-tryptophan dimethylallyl transferase (DMAT synthase). It has been isolated by the group of Floss and these investigators also purified it to homogeneity and performed the first characterisations (Heinstein et al., 1971 Lee et ai, 1976). Initially, the enzyme was reported to be a monomeric... 

Lee, S.L., Floss, H.G. and Heinstein, P. (1976) Purification and properties of dimethylallylpyrophosphate tryptopharm dimethylallyl transferase, the first enzyme of ergot alkaloid biosynthesis in Claviceps. sp. SD 58. Arch Biochem Biophys., 77, 84-94. 

Ellis, B. E., and S. A. Brown Isolation of Dimethylallylpyrophosphate Umbelli-ferone Dimethylallyl Transferase from Ruta graveolens. Canad. J. Biochem. 52, 734... 

Another derivative is the zeatin. Zeatin belongs to the plant-growth hormones, cytokinins. Cytokinins are adenine derivatives with isoprenoid side chain [108, 109]. The rate-limiting step of tra/is-zeatin biosynthesis is catalyzed by adenylate-isopentenyltransferase (cytokinin synthase) [110]. The plant enzyme (EC 2.5.1.27) utilize ADP, which is different from the microbial one (EC 2.5.1.112) which recognizes ATP/or ADP [111, 112]. Afterward, the adenylate-isopentenyl converts to the active tra/is-zeatin by a single hydroxylation reaction (cytokinin tra/is-hydroxylase). On the opposite site, cis-zeatin biosynthesis starts with a different transferase, tRNA-dimethylallyl-transferase (EC 2.5.1.75) [113-116], which mediates the transfer of an isopentenyl group of DMAPP to adenine of tRNAs. 

Consequences for the so-called segregation model ofisoprenoid biosynthesis The presence of such alternative pathways as outlined above could provide a further explanation for the observations obtained with the HMGR inhibitor mevinolin, viz. its great efficiency in blocking cytoplasmic sterol biosynthesis, but its low efficiency to affect ubiquinone biosynthesis, and its practically complete inefficiency to block the accumulation of isoprenoids and prenyl lipids of the chloroplast (summarized and discussed earlier 14, 17]. It could also explain the puzzling observation that, in contrast to phytosterols, prenyl-subsituted coumarins were not labeled from [2- ]acetate or [2- C]MVA in elicitor-treated cultures oiAmnd majus, suggesting that the DMAPP needed for the (plastid ) umbelliferone dimethylallyl transferases (EC 2.5.1.3) is formed from IPP synthesized de novo within the plastid, rather than from IPP imported into the plastid from the cytosol [86]. Elicitor-induced inhibition of phytosterol biosynthesis should occur by specific inhibition of one of the enz)ones on the c) osolic pathway from MVA to DMAPP. 

This enzyme [EC 2.5.1.1] (also referred to as prenyl-transferase and geranyl-diphosphate synthase) catalyzes the reaction of dimethylallyl diphosphate and isopen-tenyl diphosphate to produce geranyl diphosphate and pyrophosphate (or, diphosphate). The enzyme will not accept larger prenyl diphosphates as substrates. 

These isomeric C5 units condense to form a Cjq compound isopentenyl pyrophosphate attacks an allylic carbonium ion formed from dimethylallyl pyrophosphate to yield geranylpyrophosphate (Figure 26.9). The same kind of reaction takes place again geranyl pyrophosphate is converted into an allylic carbonium ion, which is attacked by isopentenyl pyrophosphate. The resulting C15 compound is called farnesylpyrophosphate. The same enzyme, geranyl transferase, catalyzes each of these condensations. 

Steroids are members of a large class of lipid compounds called terpenes. Using acetate as a starting material, a variety of organisms produce terpenes by essentially the same biosynthetic scheme (Fig. 8). The self-condensation of two molecules of acetyl coenzyme A (CoA) forms acetoacetyl CoA. Condensation of acetoacetyl CoA with a third molecule of acetyl CoA, then followed by an NADPH-mediated reduction of the thioester moiety produces mevalonic acid [150-97-0] (72). Phosphorylation of (72) followed by concomitant decarboxylation and dehydration processes produce isopentenyl pyrophosphate. Isopentenyl pyrophosphate isomerase establishes an equilibrium between isopentenyl pyrophosphate and 3,3-dimethylallyl pyrophosphate (73). The head-to-tail addition of these isoprene units forms geranyl pyrophosphate. The addition of another isopentenyl pyrophosphate unit results in the sesquiterpene (C15) famesyl pyrophosphate (74). Both of these head-to-tail additions are catalyzed by prenyl transferase. Squalene synthetase catalyzes the head-to-head addition of two achiral molecules of famesyl pyrophosphate, through a chiral cyclopropane intermediate, to form the achiral triterpene, squalene (75). 

All carotenoids are derived from the isoprenoid or terpenoid pathway. From prenyl diphosphates of different chain lengths, specific routes branch off into various terpenoid end products. The prenyl diphosphates are formed by different prenyl transferases after isomerization of IPP to DMAPP by successive T-4 condensations with IPP molecules. Condensation of one molecule of dimethylallyl diphosphate (DMADP) and three molecules of isopentyl diphosphate (IDP) produces the diter-pene geranylgeranyl diphosphate (GGDP) that forms one-half of all C40 carotenoids. The head-to-head condensation of two GGDP molecules results in the first colorless carotenoid, phytoene. Phytoene synthesis is the first committed step in C40 carotenoid biosynthesis (Britton et al. 1998, Sandmann 2001). 

A problem in the use of dimethylallyl pyrophosphate (3) is its instability. In a study of this problem the half-life of this substance was examined over a range of pH values and temperatures. Both cis- and trans-prenyl pyrophosphates (4 n = 0, 1, or 2) occur in Pinus radiata. Their biosynthesis from [2- C,3R,4K- H]-mevalonic acid proceeded with retention of tritium whereas with [2- C,3K,4S- HJmevalonic acid tritium was lost [except in the case of isopentenyl pyrophosphate (4 n = 0)]. The authors suggest that since they could not detect an isomerase, there may be a cis- and a trans-prenyl transferase both of which eliminate the label derived from [4S- H]mevalonic acid. However, compart-mentalization may have resulted in the isomerase not being available to the administered monoterpenoids, although it may act on geranyl pyrophosphate formed in situ. The absence of 6-cis-famesol derivatives tends to support this idea. Further work on this system again produced no evidence for isomerization or metabolism of [l- H]nerol pyrophosphate to 2-rrans-6-c(s-farnesyl pyrophosphate. 

The enzyme A -isopentenyl pyrophosphate tRNA A -isopentenyl transferase has been further characterized. It has a molecular weight of about 55 000, needs dimethylallyl pyrophosphate (3), and is highly stereospecific in its action. The tertiary structure of the tRNA is necessary before a reaction will occur, and the enzyme then modifies the adenosine unit adjacent to the 3 -end of the anticodon. 

The alkaloids melicopicine from Melicope fareana [86], acronycine from Acrony-chia baueri [87, 88], and rutacridone from R. graveolens (Rutaceae) typify some of the structural variety that may then ensue. For instance, radioactivity biosynthetic studies on R. graveolens, using [1- H]DMAPP (dimethylallyl diphosphate), demonstrated that 1,3-dihydroxy-A-methylacridone reacted with DMAPP upon mediation of a monoprenyl aryl transferase. The formed prenylated acridone glycocitrine-H in turn cyclized to give the dihydrofuran portion of rutacridone. Compounds 21 and 22 are hypothetical intermediates (Figure 6.18) [89]. 

Dimethylallyl pyrophosphate is the substrate for the first prenyl transferase reaction, and it is condensed with a molecule of isopentenyl pyrophosphate to give the Cio compound geranyl pyrophosphate. The addition of a second isopentenyl pyrophosphate unit produces the compound farae-syl pyrophosphate. The condensation of two molecules of famesyl pyrophosphate results in formation of the C30 compound squalene, the precursor of cholesterol and other sterols. The C20 compound, GGPP, is formed from... 

In 1973, Allen demonstrated that the monoprenylated cyclic dipeptide cyclo-L-alanyl-2-(l,l,-dimethylallyl)-L-tryptophan was incorporated into echinulin in cultures of Aspergillus amstelodami [56]. In this study, the reverse prenyiated cyclic dipeptide was doubly labeled with and by treating a partially purified prenyl transferase obtained from Aspergillus amstelodami with cyclo-L-Ala-L-[3- C]-Trp and [l- H]-3-methyl-2-butenyl 1-pyrophosphate (Scheme 33)... 

Information on the origin of angular furanocoumarins has not been plentiful, but much more is known about the more widely distributed linear isomers. In brief, the committed step in the pathway is the condensation of mevalonate-derived dimethylallyl pyrophosphate (DMAPP) at position 6 of umbelliferone to yield 7-dimethylsuberosin (Scheme 5). A transferase mediating this step was identified and characterized in Ruta graveolens 3 o 3 d evidence was obtained that it is a chloroplast enzyme.Cyclization... 

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