Phytoene desaturation reactions

A series of desaturation reactions convert phytoene to i -carotene and then to lycopene, the important red pigment in tomatoes. In pepper, lycopene undergoes a cyclization reaction on both ends by lycopene P-cyclase, thus producing P-carotene (Fig. 8.2) [25]. Beta-carotene is then converted to -cryptoxanthin, zeaxanthin. 

Biosynthesis and Metabolism.—Pathways and Reactions. Two reviews of carotenoid biosynthesis discuss, respectively, the early steps and the later reactions." The former paper deals with the mechanism of formation of phytoene and the series of desaturation reactions by which phytoene is converted into lycopene, and also describes in detail the biosynthesis of bacterial C30 carotenoids. The second paper" presents details of the mechanism and stereochemistry of cyclization and the other reactions that involve the carotenoid C-1 —C-2 double bond and the later modifications, especially the introduction of oxygen functions. 

The biosynthesis of carotenoids in plants has been reviewed extensively in recent years and is only briefly described here (Britton, 1988 Bartley and Scolnik, 1994 Sandmann, 1994). The committed step to carotenoid synthesis is the formation of the first compound phytoene by the head-to-head condensation of two molecules of GGDP by phytoene synthase. Phytoene is subjected to a series of four sequential desaturation reactions, by two separate enzymes to yield lycopene, which has eleven conjugated double bonds. Lycopene is then cyclized to /3-carotene by two /3-cyclizations or to a-carotene... 

Subsequent desaturation reactions lengthen the conjugated double-bond system to produce neurosporene or lycopene. Two completely unrelated types of phytoene desaturases exist. The enzyme found in bacteria (except cyanobacteria) and in fungi catalyzes the entire four-step desaturation process of phytoene to lycopene (Schmidt-Dannert 2000). The plant-type phytoene desaturase from cyanobacteria, algae, and plants carries out a two-step desaturation reaction with different -carotene stereoisomers as reaction products (Britton et al. 1998, Schmidt-Dannert 2000). 

Owing to the similarity of desaturation reactions catalyzed by PDS or ZDS, differentiation in the plant is not easy to detect. Most of the herbicidal inhibitors probably inhibit both, although to a different extent [6j. If strong inhibition of PDS has taken place with accumulation of phytoene, then the compound s ability to inhibit ZDS cannot be seen. Figure 4.1.2 shows that the commercial products primarily inhibit PDS [6, 8, 27-29]. Cell-free studies exemplified by norflurazon and fluridone have shown them to act as reversible noncompetitive inhibitors of PDS [27]. Other PDS active structures are shown below in Table 4.1.2 and in Section 4.1.4.10. 

In higher plants, fimgi, and some bacteria the predominant isomer of phytoene is the 15-cis compound (Section V,B,2). However, the more unsaturated carotenes in these same organisms are the all-trans isomers. This means that the pathway of phytoene desaturation in these organisms also involves an isomerization reaction. This isomerization occurs in an early stage of the pathway of phytoene desaturation, but the exact step at which it occurs may not be the same in all cases. In higher plants only traces of trans-... 

Although the pathway of phytoene desaturation is well established, little is known concerning the enzymes involved. The enzyme system from tomato fruit plastids which converts phytoene to lycopene is extractable with phosphate buffer after preparation of an acetone powder. Whether this enzyme system consists of individual enzymes or an enzyme complex is not known. It is clear, though, that an isomerase is also involved, as well as the enzymes that bring about the desaturation reactions. There is evidence for a multienzyme complex inP. blakesleeanus (Aragon et al., 1976). In this organism the... 

FIGURE 63.1 Starting with mevalonate, carotenoids are biosynthesized by a special branch of the terpenoid pathway. The first C-40 hydrocarbon unit formed is phytoene, a carotenoid with three conjugated double bonds, which then is enzymatically desaturated to successively yield (3-carotene, neurosporene, and lycopene. Other carotenoids such as (3-carotene and oxocarotenoids are produced from lycopene following cyclization and hydroxylation reactions. Thus, lycopene is a central molecule in the biosynthesis pathway of carotenoids. 

Norfluorazon is an often-used inhibitor of carotenoid biosynthesis, interfering with the step ofphytoene desaturation (Sandmann, 1994). Norflurazon treatment of mustard seedlings showed that PS II assembly is more sensitive to reduced levels of carotenoids than PS I assembly (Markgraf and Oelmiiller, 1991). The assembly of D1 into the PS II reaction center appears to be dependent on /3-carotene. Upon treatment of Chlamydomonas reinhardtii with phytoene desaturase inhibitors, D1 degraded during photoinhibition caimot be replaced (Trebst and Depka, 1997). 

Several phytoene desaturases are known that differ in the number of desaturation steps and in their structures. Among them, the bacteriaPfungal type is encoded by crti, and the cyanobacterial/algal/plant type is encoded by crtP or pds. Phytoene desaturase converts phytoene to -carotene with phytofluene as an intermediate. The reaction is stimulated by NAD, NADP, and oxygen. 

The desaturase (and following cyclase) enzymes are integral membrane proteins that can be solubilized by the detergent CHAPS. Enzymatic activity is lost when the enzymes are solubilized, but it is restored when reconstituted in proteoliposomes (Britton, 1993). In the first phase of desaturation, the conversion of (15Z)-phytoene into (15Z)- -carotene occurred in the dark, with oxygen as an essential requirement. The reaction was independent of cofactors such as NADP and NADPH. (15Z)- -Carotene could not be desaturated further, but when this compound was isomerized by light, the isomeric mixture formed underwent further desaturation to produce lycopene, not... 

Daffodil chromoplasts (Narcissus pseudonarcissus L.) convert I I-phytoene in more desaturated and cyclic carotenes. Recently, we succeeded in the dissection in vitro of these membrane bound reactions into three independent segments, the desaturation of phytoene to -carotene, the desaturation of... 

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