Mevalonic- 3-phosphate 5-pyrophosphate

Bhat, C.S. and Ramasarma, T., Effect of phenyl and phenolic acids on mevalonate-5-phosphate kinase and mevalonate-5-pyrophosphate decarboxylase of the rat brain, J. Neurochem., 32, 1531, 1979. 

Shama Bhat, C. Ramasarma, T. Inhibition of rat liver mevalonate pyrophosphate decarboxylase and mevalonate phosphate kinase by phenyl and phenolic compounds. Biochem. J., 181, 143-151 (1979)... 

The steps required to convert mevalonic acid to the active-isoprenoid intermediate have been worked out with some assurance. The initial step involves the phosphorylation of mevalonic acid to mevalonic acid-5-phosphate by an enzyme called mevalonic kinase. This enzyme was found in yeast by Tchen (1958). The properties of the mevalonic kinase of liver have been described in detail by Levy and PopjAK (1960). The kinase is inhibited by p-chloromercuribenzoate but not by iodoacetamide. The enzyme requires Mg++, Mn++, or Ca++ and ATP or inosine triphosphate. The kinase is specific for the (+) form of mevalonic acid. Mevalonic acid-5-phosphate is phosphorylated further to give mevalonic acid-5-pyrophos-phate (de Waard and Popjak, 1959 Henning et al. 1959). The purified enzyme (Bloch et al., 1959) requires a divalent metal ion for activity (Mg++ is preferable) and has no pronounced pH optimum. Mevalonic acid pyrophosphate then undergoes simultaneous dehydration and decarboxylation to yield isopentenylpyro-phosphate (Lynen et al., 1958 Chaykin et al., 1958). The enzyme concerned with the dehydration and decarboxylation has been purified (Bloch et al., 1959) and shown to have a pH optimum between 5.5 and 7.4 and to require a divalent metal ion (Mg++, Mn++, Fe++ or Co++). The series of reactions in which mevalonate is converted to isopentenylpyrophosphate is outlined in Figure 6. Brodie et al. (1963) have established a new pathway for the biosynthesis of mevalonic acid from malonyl CoA. The importance of this particular pathway in the synthesis of sterols is still unknown. 

In keeping with its biogenetic origin m three molecules of acetic acid mevalonic acid has six carbon atoms The conversion of mevalonate to isopentenyl pyrophosphate involves loss of the extra carbon as carbon dioxide First the alcohol hydroxyl groups of mevalonate are converted to phosphate ester functions—they are enzymatically phosphorylated with introduction of a simple phosphate at the tertiary site and a pyrophosphate at the primary site Decarboxylation m concert with loss of the terti ary phosphate introduces a carbon-carbon double bond and gives isopentenyl pyrophos phate the fundamental building block for formation of isoprenoid natural products... 

The terpenoid precursor isopentenyl diphosphate, formerly called isopentenyl pyrophosphate and abbreviated IPP, is biosynthesized by two different pathways depending on the organism and the structure of the final product. In animals and higher plants, sesquiterpenoids and triterpenoids arise primarily from the mevalonate pathway, whereas monoterpenoids, diterpenoids, and tetraterpenoids are biosynthesized by the 1-deoxyxylulose 5-phosphate (DXP) pathway. In bacteria,... 

The CPPase substrate DMAPP (15) is formed from isopentenyl pyrophosphate (IPP) (14) via the IPP isomerase reaction. It had been assumed that IPP was generated only via mevalonic acid (12) (Fig. 2), but Rohmer discovered another route, 2-C-methyl-D-erythritol 4-phosphate (13) (MEP) pathway (Fig. 2) [22, 23]. A key step in the MEP pathway is the reaction catalyzed by 1-deoxy-D-xylulose 5-phosphate synthase (DXS), which combines hydroxyethyl thiamine pyrophosphate (hydroxyethyl TPP) generated from pyruvic acid (17) and TPP with glyceral-dehyde 3-phosphate (18) to yield 1-deoxy-D-xylulose 5-phosphate (19) containing five carbons. The mevalonate pathway operates in the cytosol of plants and animals, whereas the MEP pathway is present in the plastid of plants or in eubacteria [24-27]. 

In pepper as in many plants, there are two sources of isoprene monomers the mevalonic acid pathway and the plastidal pool from pymvate and glyceraldehyde-3-phosphate [26], Pepper carotenoid biosynthesis uses the plastidal pathway for the isopentyl pyrophosphate monomers and the resident terpenoid synthases and transferases [27], Using the 5-carbon isoprene pool, the prenyl transferases sequentially... 

Skilleter, D.N. Kekwick, R.G.O. The enzymes forming isopentenyl pyrophosphate from 5-phosphomevalonate (mevalonate 5-phosphate) in the latex of Hevea brasiliensis. Biochem. J., 124, 407-417 (1971)... 

Mevalonic acid lactone. Mevalonic acid 5-phosphate, Mevalonic acid 5-pyrophosphate... 

Mevalonic acid 5-pyrophosphate [1492-08-6] M 258,1, Purified by ion-exchange chromatography on Dowex-1 formate [Bloch et al. JBC 234 2595 7959], DEAE-cellulose [Skilletar and Kekwick, AB 20 171 7967], on by paper chromatography [Rogers et al. BJ99 381 7966]. Likely impurities are ATP and mevalonic acid phosphate. Stored as a dry powder or as a slightly alkaline (pH 7-9) soln at -20°. 

Elimination usually involves loss of a proton together with a nucleophilic group such as -OH, -NH3+, phosphate, or pyrophosphate. However, as in Eq. 13-18, step c, electrophilic groups such as -COO-can replace the proton. Another example is the conversion of mevalonic acid-5-pyrophosphate to isopentenyl pyrophosphate (Eq. 13-19) This is a key reaction in the biosynthesis of isoprenoid compounds such as cholesterol and vitamin A (Chapter 22). The phosphate ester formed in step a is a probable intermediate and the reaction probably involves a carbo-cationic intermediate generated by the loss of phosphate prior to the decarboxylation. 

If sterol content and conformation are so important for membrane stability, we should study the biosynthesis of sterols (Figure 3). The first enzyme in terpenoid biosynthesis is the 3-Hydroxy-3-Methyl-Glutary1-Coenzyme A-reductase (HMG-CoA-reductase) that catalyzes the synthesis of mevalonate. Two phosphorylations and decarboxylation of mevalonate lead to isopentenylpyrophosphate, the basic C -unit in sterol synthesis. Isopentenylpyrophosphate reacts with its isomer, the dimethylally1-pyrophosphate, in a head/tail-reaction to geranyl-pyrophosphate reaction with another C -unit leads to farnesyl-pyro-phosphate, that dimerizes in a tail/tail-reaction to squalene. After expoxidation of its A -double bond, squalene cyclizes to lano-... 

O Reaction of mevalonic acid with three equivalents of ATP converts one hydroxy group to a pyrophosphate group and the other to a phosphate group. 

Figure 11 Biosynthesis of isoprenoid type cofactors. 18, Heme a 39, pyridoxal 5 -phosphate 43, 1-deoxy-D-xylulose 5-phosphate 46, thiamine pyrophosphate 83, acetyl-CoA 84, (S)-3-hydroxy-3-methylglutaryl-CoA 85, mevalonate 86, isopentenyl diphosphate (IPP) 87, dimethylallyl diphosphate (DMAPP) 88, pyruvate 89, D-glyceraldehyde 3-phosphate 90, 2C-methyl-D-erythritol 4-phosphate 91, 2C-methyl-erythritol 2,4-cyclodiphosphate 92, 1-hydroxy-2-methyl-2-( )-butenyl 4-diphosphate 93, polyprenyl diphosphate 94, cholecalciferol 95, fS-carotene 96, retinol 97, ubiquinone 98, menaquinone 99, a-tocopherol.

The universal precursors to terpenoids, the C5-compounds dimethylallyl pyrophosphate (DMAPP) and isopentenyl pyrophosphate (IPP), originate from two pathways in plants (Fig. 1). The mevalonate (MEV) pathway is well described in many eukaryotic organisms. This pathway is present in the cy-tosol/endoplasmic reticulum of plants. More recently, another pathway has been described, the 2C-methyl-D-erythritol-4-phosphate (MEP) pathway, which is found in the plastids of plants (19). The localization of the different pathways and the plastid-directing transit peptides found in hemi-TPS, mono-TPS, and di-TPS, but not in sesqui-TPS, result in the production of terpenoids from at least two different precursors pools. 

The natural precursors of any flavourings of this group are isopentenylpyrophosphate (IPP) and dimethylallyl pyrophosphate (DMAPP). In the classic pathway these are synthesised from three molecules acetyl/malonyl-CoA via mevalonic acid (MA). Recently a second pathway of the isoprenoid biosynthesis has been detected, in which IPP and DMAPP are synthesised from pyruvate and glyceraldehyde 3-phosphate via deoxyxylulose (DOX) 5-phosphate [322-324],... 

Isoprenyl pyrophosphates are synthesized by successive phosphorylation of mevalonate with ATP to yield the 5-monophosphate, 5-pyrophosphate, and 5-pyrophospho-3-monophospho derivatives. This last compound is very unstable and loses the 3-phosphate and the carboxyl group to yield isopentenyl pyrophosphate (IPPP), which is iso-merized to 3,3-dimethylallyl pyrophosphate (DMAPP). These reactions, catalyzed by cytosolic enzymes, are shown in Figure 19-13. 

The isopentenyl pyrophosphate and the dimethylallyl pyrophosphate precursors to the octaprenyl moiety are derived from 1-deoxy-D-xylulose-5-phosphate rather than from mevalonic acid (see also thiamin and pyridoxal sections) [183, 184]. 

Terpenoids do not necessarily contain exact multiples of five carbons and allowance has to be made for the loss or addition of one or more fragments and possible molecular rearrangements during biosynthesis. In reality the terpenoids are biosynthesized from acetate units derived from the primary metabolism of fatty acids, carbohydrates and some amino acids (see Fig. 2.10). Acetate has been shown to be the sole primary precursor of the terpenoid cholesterol. The major route for terpenoid biosynthesis, the mevalonate pathway, is summarized in Fig. 2.16. Acetyl-CoA is involved in the generation of the C6 mevalonate unit, a process that involves reduction by NADPH. Subsequent decarboxylation during phosphorylation (i.e. addition of phosphate) in the presence of ATP yields the fundamental isoprenoid unit, isopentenyl pyrophosphate (IPP), from which the terpenoids are synthesized by enzymatic condensation reactions. Recently, an alternative pathway has been discovered for the formation of IPP in various eubacteria and plants, which involves the condensation of glyceraldehyde 3-phosphate and pyruvate to form the intermediate 1-deoxy-D-xylulose 5-phosphate (Fig. 2.16 e.g. Eisenreich et al. 1998). We consider some of the more common examples of the main classes of terpenoids below. 

Most successful attempts to isolate the enzymes involved in terpene biosynthesis have come from these early stages. Crude in vitro systems will frequently convert - [2- C]mevalonic acid (1) into its phosphate (2) and pyrophosphate (3), isopentenyl pyrophosphate (4), and dimethylallyl pyrophosphate (5). However, only traces of radioactivity are recovered from the prenol pyrophosphates (6). As well as phosphorylating mevalonic acid the same enzyme, or a related one, is... 

Mevalonic acid was discovered by Folker s group at Merck, Sharpe, and Dohme. The initial isolation was based upon the fact that it acted as a growth factor, or vitamin, for a strain of bacteria [35]. Once the structure had been determined, it was apparent that the molecule might well be the isoprenoid precursor that had been sought for many years. Subsequent experiments demonstrated that the sole (or nearly so) fate of the molecule was polyisoprenoid synthesis. In examining the role of cofactors necessary for the synthesis of cholesterol from mevalonate, only ATP and NADPH were found to be required. Experiments with a solubilized preparation from yeast demonstrated that there were 3 phosphorylated intermediates that could be isolated. These were shown to be mevalonic-5-phosphate, mevalonic-5-pyrophos-phate, and isopentenyl pyrophosphate [9]. These intermediates are derived from mevalonate in a sequence of phosphorylations, and the enzymes for all reactions have been obtained in homogeneous form. These enzymes, as well as the rest that lead to the synthesis of famesyl pyrophosphate, are cytosolic proteins. 

Processes affecting the carbon-isotopic compositions of isoprenoid lipids. The isoprene carbon skeleton is indicated schematically in Figure 27. The corresponding biosynthetic reactant—equivalent in its role to acetyl-CoA—is isopentenyl pyrophosphate. As shown in Figure 29, this compound can be made by two different and fully independent pathways. The mevalonic-acid pathway was until recently thought to be the only route to isoprenoids. The deoxyxylulose-phosphate, or methylerythritol-phosphate, pathway was first discovered in Bacteria by Rohmer and coworkers (Flesch and Rohmer... 

Figure 29. Relationships h een the carbon positions in isopentenyl pyrophosphate and their sources. In the mevalonic-acid pathway, all five caibon positions in isopentenyl pyrophosphate derive from acetate and, in turn from the C-1 + C-6 and C-2 + C5 positions of glucose. In the methyierythritol-phosphate pathway, one carbon derives from the C-3 + C-4 position in glucose. Moreover, the mapping of positions from preciu ors into products of the two pathways differs sharply, as indicated by stmctures of acyclic and steroidal carbon skeletons based on the MVA (a, c) and MEP pathways (b, d).

The mechanism for converting mevalonic acid into mevaionyi phosphate is essentially an Sn2 reaction with an adenosyl pyrophosphate leaving group (Section 27.3). A second Sn2 reaction converts mevaionyi phosphate to mevaionyi pyrophosphate. ATP is an excellent phosphorylating reagent for nucleophiles because its phosphoanhydride bonds are easily broken. The reason that phosphoanhydride bonds are so easily broken is discussed in Section 27.4. 

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