Bacterial phytoene

Over-expression of bacterial phytoene synthase led to only modest increases in pigment accumulation (except in the case of chloroplast-contaiifing tissues). Attention turned to CrtI, one gene that might control flux through the entire four desaturation steps from phytoene to lycopene (discussed in Section 5.3.2.4). Only a modest increase in carotenoid content in tomatoes and a variety of changes in carotenoid composition including more P-carotene, accompanied by an overall decrease in total carotenoid content (no lycopene increase), resulted when CrtI was over-expressed under control of CaMV 35S. Apparently, the bacterial desaturase... 

A number of other studies have focused on increasing the levels of P-carotene in tomato (Table 56.6). Ectopic expression of a bacterial phytoene synthase (crtB) in a fruit-specific manner increased phytoene (2.4-fold), lycopene (1.8-fold), p-carotene (2.2-fold), and lutein levels in tomato fruit [113]. Suppression RNAi was used to downregulate DE-ETIOLATEDl (DETl), photomorphogenesis... 

Eujisawa M, Watanabe M, Choi S-K, Teramoto M, Ohyama K, Misawa N (2008) Enrichment of carotenoids in flaxseed (Linum usitatissimum) by metabolic engineering with introduction of bacterial phytoene synthase gene crtB. J Biosci Bioeng 105 636-641... 

Fortuitously, the bacterial gene product, CRTI, produces di -trans carotenoids and satisfies the stereo-chemical specificity of LYC B for all-trani substrates while also catalyzing the four desaturation steps from phytoene to lycopene. Nevertheless, over-expression of Crtl has been shown to have only a modest effect (two- to fourfold increases in tomatoes and carrots) in increasing flux through the pathway and some unexpected pleiotropic influences on activities upstream and downstream of the desaturations (reviewed by Fraser and Bramley and Giuliano °). 

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. 

Phytoene synthase is encoded by the closely related bacterial crtB and eukaryotic psy genes. The phytoene formed in the enzymatic reaction was present in both a 15-cw and all-trans isomeric configuration. The essential cofactors required were adenosine triphosphate (ATP) in combinations with either Mn + or Mg +. Phytoene synthesis was inhibited by phosphate ions and squalestatin. 

The use of hmgr-1 resulted in elevated levels of phytosterols (up to 2.4-fold), but IPP-derived isoprenoids in the plastid were unaltered. Transgenic lines containing the bacterial dxs targeted to the plastid with the tomato dxs transit sequence resulted in an increased carotenoid content (1.6-fold), with phytoene and p-carotene exhibiting the highest increases (2.4- and 2.2-fold, respectively). This result emphasizes the idea that IPP used for the carotenoid pathway is mostly obtained from the MEP pathway [24],... 

Tsuchiya T, Takaichi S, Misawa N, Maoka T, Miyashita H, Mimuro M (2005) The cyanobacterium Gloeobacter violaceus PCC 7421 uses bacterial-type phytoene desaturase in carotenoid biosynthesis. FEES Lett 579 2125-2129... 

Harada J, Nagashima KVP, Takaichi S, Misawa N, Matsuura K, Shimada K (2001) Phytoene desaturase, CrtI, of the purple photosynthetic bacterium, Rubrivivax gelatinosus, produces both neurosporene and lycopene. Plant Cell Physiol 42 1112-1118 Francis GW, Liaaen-Jensen S (1970) Bacterial carotenoids XXXIII. Carotenoids of thiorhodaceae 9. The structures of the carotenoids of the rhodopinal series. Acta Chem Scand 24 2705-2712... 

In the cyclization of phytoene [4.122] a particular stereochemistry is expected for proton and carbon addition to the C-1 double bond. [2- C]Mevalonate labels the ( )-methyl group at C-1 in lycopene [[4.127 = [4.123] = label] and the la-methyl group in zeaxanthin [[4.128 = [4.125 (see Scheme 4.28). Zeaxanthin formed in bacterial cells suspended in deuterium oxide had a 2) -deuteron. It follows that cyclization occurs as shown in Scheme 4.28 [111]. The stereochemistry in the C50 carotenoids, e.g. [4.126] implies a different manner of addition to the C-1 double bond. 

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