Zeaxanthin epoxidase

The oxidation of carotenes results in the formation of a diverse array of xanthophylls (Fig. 13.7). Zeaxanthin is synthesised from P-carotene by the hydroxylation of C-3 and C-3 of the P-rings via the mono-hydroxylated intermediate P-cryptoxanthin, a process requiring molecular oxygen in a mixed-function oxidase reaction. The gene encoding P-carotene hydroxylase (crtZ) has been cloned from a number of non-photosynthetic prokaryotes (reviewed by Armstrong, 1994) and from Arabidopsis (Sun et al, 1996). Zeaxanthin is converted to violaxanthin by zeaxanthin epoxidase which epoxidises both P-rings of zeaxanthin at the 5,6 positions (Fig. 13.7). The... 

CRTR-e = e-ring hydroxyiase CRTR-b = p-ring hydroxylase ZEP-1 = zeaxanthin epoxidase NXS = neoxanthin synthase VDE 1 = violaxanthin de-epoxidase. 

A (1996) Molecular identification of zeaxanthin epoxidase of Nicotiana plumbaginifolia, a gene involved in abscisic acid biosynthesis and corresponding to the ABA locus of Arabidopsis thaliana , EMBO J, 15, 331-42. 

The 3-hydroxyl P-rings of zeaxanthin are further oxygenated by the introduction of 5,6-epoxy moieties by zeaxanthin epoxidase (ZEP). A mono-epoxidated intermediate, antheraxanthin is produced, followed by the di-epoxy xanthophyU, violaxanthin, as shown in Figure 5.3.3B. 

Thompson, A. J., A. C. Jackson et al. (2000a). Abscisic acid biosynthesis in tomato Regulation of zeaxanthin epoxidase and 9-cw-epoxycarotenoid dioxygenase mRNAs by light/dark cycles, water stress and abscisic acid. Plant Mol. Biol. 42(6) 833-845. 

Figure 73. The carotenoid biosynthetic pathway. Enzymes are named according to the designation of their genes Ccs, capsanthin-capsorubin synthase CrtL-b, lycopene-b-cyclase CrtL-e, lycopene-e-cyclase CrtR-b, b-ring hydroxylase, CrtR-e, e-ring hydroxylase DMADP, dimethylallyl diphosphate GGDP, geranylgeranyl diphosphate Ggps, geranylgeranyl-diphosphate synthase IDP, isopentenyl diphosphate Ipi, IDP isomerase Pds, phytoene desaturase Psy, phytoene synthase Vde, violaxanthin de-epoxidase Zds, z-carotene desaturase Zep, zeaxanthin epoxidase. (From van den Berg and others 2000.)...

Fig. 8.2 Carotene and xanthophyll biosynthetic pathways in Capsicum. Isopentenyl pyrophosphate (IPP) phytoene synthase (PSY) lycopene fi-cyclase (LCYB) lycopene e-cyclase (LCYE) P-carotene hydroxylase (CrtZ-2) zeaxanthin epoxidase (Ze) and capsanthin-capsorubin synthase (CCS)...

Fig. /. Carotenoid biosynthetic pathway in plants. For clarity, only the portion of the biosynthetic pathway starting with lycopene is shown. Enzymes are indicated by numbers as follows I, /J-cyclase 2, e-cyclase 3, / -ring hydroxylase 4, e-ring hydroxylase 5, zeaxanthin epoxidase and violaxanthin de-epoxidase enzymes 6, neoxanthin forming enzyme. The location of the lutJy lut2, and aba mutations are indicated. Carotenoids that typically accumulate in photosynthetic tissues are given in capital letters.

Chlamydomonas mutants have been isolated that are deficient either in violaxanthin de-epoxidase or zeaxanthin epoxidase. Using these mutants, it could be demonstrated that only part of the non-photochemical quenching observed in Chlamydomonas is dependent on the formation of zeaxanthin (Niyogi etal., 1997a). It will be interesting to see whether the absence of xanthophyll epoxidase or de-epoxidase products has an effect on the assembly of light-harvesting complexes in these mutants. 

Fig 2. Schematic illustrating domains identified within the coding sequence of the deduced amino acid sequences for violaxanthin de-epoxidase and zeaxanthin epoxidase. (A) Violaxanthin de-epoxidase from Lactuca saliva (Bugos and Yamamoto, 1996). (B) Zeaxanthin epoxidase from Nicotianaplumbaginifolia (Marin et al., 1996), The domains for each sequence are illustrated (1- III) with the amino acid sequence spanning regions indicated below. The amino acid numbering is from the N-terminus of the mature protein. 

The carotenoid content of the mutant and wild-type plants was analyzed by HPLC to determine at what step the ABA biosynthetic pathway was impaired. It was found that zeaxanthin content accumulated in the mutant but not in the wild-type leaves. In addition, violaxanthin and neoxanthin were not detected in mutant leaves. Therefore, it was concluded that the mutant was impaired in the epoxidation of zeaxanthin. DNA sequence information from the regions flanking the Ac insertion site was then used to screen a cDNA library and a gene encoding a chloroplast imported protein was isolated and subsequently identified to be a zeaxanthin epoxidase. 

Fig. I. The xamhophyll cycle and its effects on light-harvesting and energy dissipation within the photosynthetic apparatus. The enzyme violaxanthin de-epoxidase (VDE) catalyzes the sequential conversion of violaxanthin to zeaxanthin via antheraxanthin using ascorbic acid (AA) as a reductant and generating monodehydroascorbate (MDHA). The epoxidation of zeaxanthin to violaxanthin in the reverse reaction sequence is catalyzed by the enzyme zeaxanthin epoxidase (ZE).

Chlamydomonas reinhardtii double mutant npq2/lorl lacks the P,e-carotenoids lutein and loroxanthin as well as all P,P-epoxycarotenoids derived from zeaxanthin (e g. violaxanthin and neoxanthin). Thus, the only carotenoids present in the thylakoid membranes of the npq2/lorl cells are P-carotene and zeaxanthin. The effect of these mutations and the lack of specific xanthophylls on the Chi antenna size of the photosystems was investigated [16]. In cells of the mutant strain, the Chi antenna size of PSII was substantially smaller than that of the wild type (Table 1). In contrast, the Chi antenna size of PSI was not truncated in the mutant. This analysis showed that absence of lutein, violaxanthin and neoxanthin specifically caused a smaller functional Chi antenna size for PSII but not for that of PSI. Thus, xanthophyll-bios5mthesis genes, such as lycopene e-cyclase and zeaxanthin epoxidase may be targets for a truncated Chi antenna size in PSII. 

Fig. 56.2 Carotenoid biosynthesis pathway in plants. Enzymes (with abbreviations) indicated are isopentenyl pyrophosphate isomerase (IPI), geranylgeranyl pyrophosphate synthase (GGPS), phytoene synthase (PSY), phytoene desaturase (PDS), zeta-carotene desaturase (ZDS), carotenoid isomerase (CRTISO), lycopene beta-cyclase (LCYB), lycopene epsilon-cyclase (LCYE), beta-ring carotene hydroxylase (CHXB), epsilon-ring carotene hydroxylase (CHXE), zeaxanthin epoxidase (ZEP), violaxanthin de-epoxidase (VDE), capsorubin-capsanthin synthase (CCS), neoxanthin synthase (NXS), 9-cis epoxycarotenoid dioxygenase (NCED), and carotenoid cleavage dioxygenase (CCD). (Source [101], drawn using KeGG pathway)...

Epoxidation of zeaxanthin by zeaxanthin epoxidase (ZE) would result in the production of violaxanthin via antheraxanthin. From that substrate, the enzyme neoxanthin synthase (NXS) would yield neoxanthin opening the cyclohexenyl 5-6 epoxide ring in violaxanthin [38]. Neoxanthin would be the last product of carotenoid biosynthesis in green parts of the plant, and it would derive in the abscisic acid (ABA) synthesis pathway. The accumulation of neoxanthin and violaxanthin in flowers results in wildtype yellow petals. A defective mutation in the gene encoding CRTR-B2 prevents formation of these xanthophylls, resulting in the white-flower phenotype [18]. 

Burbidge A, Grieve T, Terry C, Corlett J, Thompson A, Taylor I (1997) Structure and expression of a cDNA encoding zeaxanthin epoxidase, isolated from a wilt-related tomato Lycopersicon esculentum Mill.) library. J Exp Bot 48 1749-1750... 

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