Presqualene pyrophosphate, biosynthesis

Most of the compounds cited in this introductory section are produced in metabolic processes where the cyclopropane-containing metabolite appears to be the stable end product or secondary product with as yet unobvious metabolic function. However, this is not the case in at least two types of systems, in which cyclopropyl species are key and necessary intermediate structures in high flux metabolic pathways. The first example is the squalene (76) and phytoene (88) biosynthesis where presqualene pyrophosphate (77) and prephytoene pyrophosphate (89) are obligate cyclopropanoid intermediates in the net head-to-head condensations of two farnesyl pyrophosphate (73) or two geranylgeranyl pyrophosphate (66) molecules respectively. The second example is in plant hormone metabolism where C(3) and C(4) of the amino acid methionine are excised as the simple hormone ethylene via intermediacy of 1-aminocyclopropane-l-carboxylic acid (9). Both examples will be discussed in detail in the Section II. 

First, we will take up cyclopropyl group formation by the rearrangement of carbon skeletons via cationic intermediates encountered in various mono- and sesquiterpenes, and also examine the illudin biosynthesis where contraction of a cyclobutyl cation to a cyclopropane has been invoked. We will then discuss the head-to-head condensation of isoprenoid alcohols at the C15 or C20 level to generate the cyclopropyl intermediates, presqualene pyrophosphate and prephytoene pyrophosphate, on the way to the C30 and C40 polyene hydrocarbons, squalene and phytoene respectively. Conversion of 2,3-oxidosqualene via common intermediate protosterol cation to cycloartenol or lanosterol represents an important pathway in which the angular methyl group participates in the three-membered ring formation. The cyclopropanation outcome of this process has been carefully studied. 

In this section we analyze information about metabolic cleavage or breakdown of cyclopropane rings in three instances the biosynthesis of irregular monoterpenes, the ringopening of cycloartenol (20) derivatives, and the metabolic opening of 1-aminocyclopropane-1-carboxylic acid (ACPC) (9) by two quite distinct fragmentation routes. We will not explicitly discuss the processing of presqualene pyrophosphate (77) and prephytoene pyrophosphate (89) to squalene (76) and phytoene (88) respectively, since those transformations have already been dealt with in Section II. 

The ergosterol (45) biosynthesis in fungi and leishmania utilizes squalene (41) as starting material, which is obtained from long chain precursors like farnesyl pyrophosphate (39) and presqualene pyrophosphate (40). Epoxidation of squalene in the presence of squalene epoxidase furnishes squalene epoxide (42), which is succes-... 

The resulting cyclopropylcarbinyl pyrophosphate is reduced to squalene by NADPH. This is the head-to-head condensation of terpene biosynthesis. The absolute stereochemistry of presqualene pyrophosphate has been determined [69]. 

The mevalonate pathway in the cytosol is responsible for biosynthesis of sterols, sesquiterpenes, and triterpenoids. After conversion of mevalonic acid to isopentenyl pyrophosphate, three C5 units can be joined head to tail to produce a C15 compound, famesyl pyrophosphate. Two famesyl pyrophosphates are then united head to head to form squalene, the progenitor of the C30 isoprenoids from which sterols are derived. The plant squalene synthetase, like its mammalian homologue, is found in the ER and the reaction proceeds via a presqualene pyrophosphate intermediate (Chapter 14). In the last step prior to cyclization, squalene is converted to squalene 2,3-epoxide. 

In cholesterol biosynthesis, farnesyl pyrophosphate is converted to presqualene pyrophosphate, as follows ... 

Squalene is an intermediate in cholesterol and steroid biosynthesis. It is formed from presqualene pyrophosphate (Figure 19.21) in the walls of the endoplasmic reticulum using electrons from NADPH. In the reaction, the pyrophosphate is removed from the molecule. 

See also Presqualene Pyrophosphate, Lanosterol, Cholesterol Biosynthesis, Steroid Metabolism, Steroid Hormone Synthesis... 

In studies of the biosynthesis of presqualene alcohol with [l-0 ]famesyl pyrophosphate, exchange of oxygen did not occur. These observations indicate that the pyrophosphate group of one famesyl pyrophosphate molecule occurs in the product without change of configuration in presqualene pyrophosphate (Popjak et al., 1975). 

Fig. 23.3. Presqualene pyrophosphate and chrysanthemic acid and inhibitors for squalene biosynthesis.

The last step in the biosynthesis of phytoene is the head-to-head condensation of two molecules of GGPP (Fig. 8). In most respects, this reaction is analogous to the condensation of two molecules of famesyl pyrophosphate to form squalene in the sterol biosynthetic pathway. Studies on the latter reaction have provided a great deal of insight into the mechanism of phytoene biosynthesis and are included in the discussion below where relevant. Both reactions involve the formation of a cyclopropylcarbinyl pyrophosphate intermediate. In the biosynthesis of phytoene this compound is pre-phytoene pyrophosphate, the C40 analogue of presqualene pyrophosphate. A proposed mechanism for the formation of prephytoene pyrophosphate is shown in Fig. 9 (Beytia and Porter, 1976). Prephytoene pyrophosphate has been isolated from a Mycobacterium preparation and synthesized chemically. Both the natural and synthetic compounds have been converted to phytoene by a cell-fiee system from Mycobacterium (Altman et al., 1972). Prephytoene pyrophosphate was also formed when GGPP was incubated with yeast squalene synthetase (Qureshi et al., 1972, 1S>73), but in the pres-... 

In the biosynthesis of steroids lanosterol and cycloartenol, the building blocks DMAPP and IPP are first joined through famesyl diphosphate synthase to form geranyl pyrophosphate (GPP) and then to form FPP, presqualene pyrophosphate, squalene, and (S)-squalene-2,3-epoxide by corresponding enzymes, sequentially. The product (S)-squalene-2,3-epoxide is used to synthesize the sterols lanosterol (in animals) and cycloartenol (in plants) by lanosterol synthase and cycloartenol synthase, respectively [3]. Further, lanosterol and cycloartenol are converted into other steroids through steroidogenesis. 

The next step in understanding squalene biosynthesis depended on the isolation, from yeast and from rat liver microsomal incubations in the absence of NADPH, of a cyclopropyl compound, presqualene pyrophosphate 4.48) it is established as an intermediate in squalene biosynthesis (in micromosal preparations it is converted into squalene in the presence of NADPH) [57-60]. A reasonable mechanism for the biosynthesis of squalene 4.47) via presqualene pyrophosphate 4.48) is shown in Scheme 4.12 [60-63]. 

Most of the natural carotenoid pigments are tetraterpenes, being formed from two C20 geranylgeranyl pyrophosphate units through phytoene 4.122). The mechanism of phytoene formation has much in common with squalene biosynthesis. In particular, head-on fusion of the two C20 units occurs via prephytoene pyrophosphate 4.120), analogous to presqualene pyrophosphate 4.48) formation from two... 

Fig. 7. A mechanism for biosynthesis of botryococcene (7) and squalene (25) from presqualene pyrophosphate (24) (R = C11H19). Reproduced with the permission of J. Amer. Chem. Soc. [Ill, 2713, (1989)]...

Figure 3.31 Biosynthesis of presqualene pyrophosphate and squalene from FPP according to Blagg etal. (2002).

Unexpected results have come to light bearing on monoterpenoid biosynthesis (Chapter 1). Banthorpe s group have shown that in the formation of the thujane and camphor skeletons, activity from labelled mevalonic acid can appear predominantly in the C5 unit supposedly derived from isopentenyl pyrophosphate and only to a minor extent in the dimethylallyl pyrophosphate-derived portion. Banthorpe has also presented evidence for a chrysanthemyl intermediate, analogous to presqualene alcohol, in the biosynthesis of artemesia ketone. 

The controversy over presqualene alcohol has been resolved in favour of Rilling s second structure (5), rather than the diester proposed by Popjak or the acyclic formulation suggested by Lynen. In the biosynthesis from farnesyl pyrophosphate one hydrogen atom is lost to the medium from C-1, and when the presqualene alcohol pyrophosphate is further metabolized to squalene (6) no further hydrogen atoms are lost. Final proof of the structure came from its synthesis by three groups the indicated absolute stereochemistry was based on a correlation with trans-chrysanthemyl alcohol. This structure is now also accepted by Popjak and co-workers. Thus the conversion of farnesyl pyrophosphate into squalene may be rationalized as shown (see also ref 29). 

A series of analogues, (4) and (5), of the precursors of squalene, in which the carbinol oxygen is replaced by a methylene group, inhibited the formation of squalene from [2- C]- and [S- HaJ-MVA by rat liver squalene synthetase. These phosphonophosphates also inhibited the biosynthesis of kaurene from MVA in a cell-free system from Ricinus communis, but the corresponding phosphonates were only weakly inhibitory. The formation of labelled squalene from a mixture of [2- C]MVA and [l- H]presqualene alcohol pyrophosphate by the squalene synthetase preparation was completely inhibited by the addition of (5) to the incubation... 

Prephytoene alcohol pyrophosphate (22b) has been synthesized and identified as an intermediate in the biosynthesis of phytoene (24b). The reaction sequence shown (Scheme 2 b series) assumes that the absolute stereochemistry is the same as for presqualene alcohol. 

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