Determination of the stoichiometry and strength of binding of xanthophylls to the photosystem II light harvesting complexes

Xanthophylls have a crucial role in the structure and function of the light harvesting complexes of photosystem II (LHCII) in plants. The binding of xanthophylls to LHCII has been investigated, particularly with respect to the xanthophyll cycle carotenoids violaxanthin and zeaxanthin. It was found that most of the violaxanthin pool was loosely bound to the major complex and could be removed by mild detergent treatment. Gentle solubilization of photosystem II particles and thylakoids allowed the isolation of complexes, including a newly described oligomeric preparation, enriched in trimers, that retained all of the in vivo violaxanthin pool. It was estimated that each LHCII monomer can bind at least one violaxanthin. The extent to which different pigments can be removed from LHCII indicated that the relative strength of binding was chlorophyll b > neoxanthin > chlorophyll a > lutein > zeaxanthin > violaxanthin. The xanthophyll binding sites are of two types: internal sites binding lutein and peripheral sites binding neoxanthin and violaxanthin. In CP29, a minor LHCII, both a lutein site and the neoxanthin site can be occupied by violaxanthin. Upon activation of the violaxanthin de-epoxidase, the highest de-epoxidation state was found for the main LHCII component and the lowest for CP29, suggesting that only violaxanthin loosely bound to LHCII is available for de-epoxidation.     

Seasonal changes of violaxanthin cycle pigment de-epoxidation in wintergreen and evergreen plants

We studied carotenoids composition and the activities of the xanthophylls pigments in evergreen conifers (Abies sibirica, Juniperus communis, Picea obovata) and dwarf-shrub (Vaccinium vitis-idaea), and in wintergreen herbaceous plants (Ajuga reptans, Pyrola rotundifolia) growing near Syktyvkar (61°67(/) N 50°77(/) E). The carotenoid pool consisted mainly of following xanthophylls: lutein (70%), neoxanthin (7-10%) and a xanthophylls cycle component - violaxanthin (3-15%). Zeaxanthin and antheraxanthin were found in conifer samples collected in December-March while in other species - during all year. A direct connection between xanthophyll pigment de-epoxidation level and light energy thermal dissipation was shown only for boreal conifer species. It is proposed that zeaxanthin plays a central role in the dissipation of excess excitation energy (nonphotochemical quenching) in the antenna of photosystem II (PSII). We conclude that the increase in the extent of de-epoxidation is beneficial for the retention of PSII activity for conifers in early spring and for herbs in summer.     

Carotenoid binding sites in LHCIIb. Relative affinities towards major xanthophylls of higher plants.

The major light-harvesting complex of photosystem II can be reconstituted in vitro from its bacterially expressed apoprotein with chlorophylls a and b and neoxanthin, violaxanthin, lutein, or zeaxanthin as the only xanthophyll. Reconstitution of these one-carotenoid complexes requires low-stringency conditions during complex formation and isolation. Neoxanthin complexes (containing 30-50% of the all-trans isomer) disintegrate during electrophoresis, exhibit a largely reduced resistance against proteolytic attack; in addition, energy transfer from Chl b to Chl a is easily disrupted at elevated temperature. Complexes reconstituted in the presence of either zeaxanthin or lutein contain nearly two xanthophylls per 12 chlorophylls and are more resistant against trypsin. Lutein-LHCIIb also exhibits an intermediate maintenance of energy transfer at higher temperature. Violaxanthin complexes approach a xanthophyll/12 chlorophyll ratio of 3, similar to the ratio in recombinant LHCIIb containing all xanthophylls. On the other hand, violaxanthin-LHCIIb exhibits a low thermal stability like neoxanthin complexes, but an intermediate accessibility towards trypsin, similar to lutein-LHCIIb and zeaxanthin-LHCIIb. Binary competition experiments were performed with two xanthophylls at varying ratios in the reconstitution. Analysis of the xanthophyll contents in the reconstitution products yielded information about relative carotenoid affinities of three assumed binding sites. In lutein/neoxanthin competition experiments, two binding sites showed a strong preference (> 200-fold) for lutein, whereas the third binding site had a higher affinity (25-fold) to neoxanthin. Competition between lutein and violaxanthin gave a similar result, although the specificities were lower: two binding sites have a 36-fold preference for lutein and one has a fivefold preference for violaxanthin. The lowest selectivity was between lutein and zeaxanthin: two binding sites had a fivefold higher affinity for lutein and one has a threefold higher affinity to zeaxanthin.     

Leaf Xanthophyll content and composition in sun and shade determined by HPLC

As a part of our investigations to test the hypothesis that zeaxanthin formed by reversible de-epoxidation of violaxanthin serves to dissipate any excessive and potentially harmful excitation energy we determined the influence of light climate on the size of the xanthophyll cycle pool (violaxanthin + antheraxanthin + zeaxanthin) in leaves of a number of species of higher plants. The maximum amount of zeaxanthin that can be formed by de-epoxidation of violaxanthin and antheraxanthin is determined by the pool size of the xanthophyll cycle. To quantitate the individual leaf carotenoids a rapid, sensitive and accurate HPLC method was developed using a non-endcapped Zorbax ODS column, giving baseline separation of lutein and zeaxanthin as well as of other carotenoids and Chl a and b.The size of the xanthophyll cycle pool, both on a basis of light-intercepting leaf area and of light-harvesting chlorophyll, was ca. four times greater in sun-grown leaves of a group of ten sun tolerant species than in shade-grown leaves in a group of nine shade tolerant species. In contrast there were no marked or consistent differences between the two groups in the content of the other major leaf xanthophylls, lutein and neoxanthin. Also, in each of four species examined the xanthophyll pool size increased with an increase in the amount of light available during leaf development whereas there was little change in the content of the other xanthophylls. However, the -carotene/-carotene ratio decreased and little or no -carotene was detected in sun-grown leaves. Among shade-grown leaves the -carotene/-carotene ratio was considerably higher in species deemed to be umbrophilic than in species deemed to be heliophilic.The percentage of the xanthophyll cycle pool present as violaxanthin (di-epoxy-zeaxanthin) at solar noon was 96–100% for shade-grown plants and 4–53% for sun-grown plants with zeaxanthin accounting for most of the balance. The percentage of zeaxanthin in leaves exposed to midday solar radiation was higher in those with low than in those with high photosynthetic capacity.The results are consistent with the hypothesis that the xanthophyll cycle is involved in the regulation of energy dissipation in the pigment bed, thereby preventing a buildup of excessive excitation energy at the reaction centers.Abbreviations A antheraxanthin - C -carotene - C -carotene - EPS epoxidation state (V+0.5A)/(V+A+Z) - L lutein - N neoxanthin - PFD photon flux density - V violaxanthin - Z zeaxanthin C.I.W.-D.P.B. Publiation No. 1035     

Relationships between carotenoid composition and growth habit in British plant species

The pigment composition of leaves from a number of different plant species collected from field sites in the region of Sheffield, UK, have been compared using high-performance liquid chromatography. Expression of pigment content per unit leaf area was dominated by variation in the total leaf chlorophyll. Neither chlorophyll per unit area nor the chlorophyll a/b ratio were found to be correlated with the habitat from which the plants originated. When the amounts of different carotenoids were expressed relative to the total carotenoid pool, it was found that whilst neither total carotene (α- +β-carotene) nor neoxanthin correlated with ability to grow in shade, the leaf content of both lutein and the total xanthophyll cycle carotenoids (zeaxanthin, anther-axanthin and violaxanthin) did, with lutein content being high in shade species and xanthophyll cycle intermediates low. There was a strong negative correlation between the relative amounts of each of these groups of carotenoids. The ratio of lutein to xanthophyll cycle carotenoids was strongly correlated to an index of shade tolerance.     

Carotenoid composition in sun and shade leaves of plants with different life forms

The carotenoid composition of sun leaves of nine species of annual crop plants (some with several varieties) was compared with sun and shade leaves of several other groups of plants, among those sun and shade leaves of several species of perennial shrubs and vines and deep-shade leaves of seven rainforest species. All sun leaves contained considerably greater amounts of the components of the xanthophyll cycle violaxanthin, antheraxanthin and zeaxanthin as well as of β-carotene than the shade leaves, as had previously been reported for a variety of other species by Thayer & Björkman (Photosynthesis Research, 1990, 23, 331–343). Therefore, high light specifically stimulated β,β-carotenoid synthesis. The sun leaves of these crop species did not contain α-carotene which was, however, present in large amounts in all shade leaves and in smaller amounts in sun leaves of three of the four species of perennial shrubs and vines. There was no difference in neoxanthin content on a chlorophyll basis between sun and shade leaves, and there was no consistent general difference in the lutein content between all sun and all shade leaves. The zeaxanthin (and antheraxanthin) content at peak irradiance and the xanthophyll cycle pool size were compared for sun leaves from the different groups of plants with different life forms and different metabolic activities. When growing in full sunlight the annual crop species and a perennial mesophyte had high rates of photosynthesis whereas the perennial shrubs and vines had relatively low photosynthesis rates. More zeaxanthin (and antheraxanthin) were accumulated at noon in full sunlight in those species with the lower photosynthesis rates. However, it was not such that those species also possessed the larger pools of violaxanthin + antheraxanthin + zeaxanthin. Instead, the xanthophyll cycle pools of sun leaves of the annual crop species and the perennial mesophyte were not smaller, and were even possibly larger, than those of sun leaves of the perennial shrubs and vines with low photosynthesis rates. This was so in spite of the fact that the crop species experienced much lesser degrees of excessive light at full sun than the shrubs and vines. Thus, many of the crop species converted only about 30–50% of their xanthophyll cycle pool to zeaxanthin at noon, whereas the shrubs and vines typically converted more than 80% of their pool into zeaxanthin. The crop species also had larger pools of β-carotene than the shrubs and vines but smaller pools of lutein than the majority of the latter species.     

Different roles of alpha- and beta-branch xanthophylls in photosystem assembly and photoprotection

Xanthophylls (oxygenated carotenoids) are essential components of the plant photosynthetic apparatus, where they act in photosystem assembly, light harvesting, and photoprotection. Nevertheless, the specific function of individual xanthophyll species awaits complete elucidation. In this work, we analyze the photosynthetic phenotypes of two newly isolated Arabidopsis mutants in carotenoid biosynthesis containing exclusively alpha-branch (chy1chy2lut5) or beta-branch (chy1chy2lut2) xanthophylls. Both mutants show complete lack of qE, the rapidly reversible component of nonphotochemical quenching, and high levels of photoinhibition and lipid peroxidation under photooxidative stress. Both mutants are much more photosensitive than npq1lut2, which contains high levels of viola- and neoxanthin and a higher stoichiometry of light-harvesting proteins with respect to photosystem II core complexes, suggesting that the content in light-harvesting complexes plays an important role in photoprotection. In addition, chy1chy2lut5, which has lutein as the only xanthophyll, shows unprecedented photosensitivity even in low light conditions, reduced electron transport rate, enhanced photobleaching of isolated LHCII complexes, and a selective loss of CP26 with respect to chy1chy2lut2, highlighting a specific role of beta-branch xanthophylls in photoprotection and in qE mechanism. The stronger photosystem II photoinhibition of both mutants correlates with the higher rate of singlet oxygen production from thylakoids and isolated light-harvesting complexes, whereas carotenoid composition of photosystem II core complex was not influential. In depth analysis of the mutant phenotypes suggests that alpha-branch (lutein) and beta-branch (zeaxanthin, violaxanthin, and neoxanthin) xanthophylls have distinct and complementary roles in antenna protein assembly and in the mechanisms of photoprotection.     

Carotenoid composition in Zea mays developed at sub-optimal temperature and different light intensities

The content and composition of pigments were examined in the third leaf of Zea mays L. plants grown under controlled environment at near-optimal temperature (24°C) or sub-optimal temperature (14°C) at a light intensity of either 200 or 600 μmol m^?2 s^?1. Compared to leaves grown at 24°C, leaves grown at 14°C showed a large reduction in the chlorophyll (Chl) content, a marked decrease in the Chl a/b ratio, and a large increase in the ratio of total carotenoids/Chl a+b. Leaves grown at 14°C showed a much lower content of β-carotene than leaves grown at 24°C, while the content of the carotenoids of the xanthophyll cycle (violaxanthin [V] + antheraxanthin [A] + zeaxanthin [Z]) was markedly higher in the former leaves as compared to the latter leaves; neoxanthin and lutein were affected by the growth temperature to a much lesser extent. The xanthophylls/β-carotene ratio was about three times higher in leaves grown at 14°C as compared to leaves grown at 24°C. On a chlorophyll basis, the two types of leaves hardly differed in their level of β-carotene, while the levels of the xanthophylls (including lutein and neoxanthin) were higher in 14°C-grown leaves as compared to 24°C-grown leaves. In leaves grown at 14°C, 40 and 56% of the V+A+Z pool was in the form of zeaxanthin at low light intensity and high light intensity, respectively. Only trace amounts of zeaxanthin, if any, were present in leaves grown at 24°C. The changes in the pigment composition induced by growth at sub-optimal temperature were more pronounced at a light intensity of 600 as compared to 200 μmol m^?2 s^?1. In the given range, the light intensity slightly affected the composition of pigments in leaves grown at 24°C. The physiological significance of the modifications to the pigment composition induced by growth at sub-optimal temperature is discussed.     

Characterization of the Xanthophyll Cycle and Other Photosynthetic Pigment Changes Induced by Iron Deficiency in Sugar Beet (Beta vulgaris L.)

In this work we characterize the changes induced by iron deficiency in the pigment composition of sugar beet (Beta vulgaris L.) leaves. When sugar beet plants were grown hydroponically under limited iron supply, neoxanthin and β-carotene decreased concomitantly with chlorophyll a, whereas lutein and the carotenoids within the xanthophyll cycle were less affected. Iron deficiency caused major increases in the lutein/chlorophyll a and xanthophyll cycle pigments/chlorophyll a molar ratios. Xanthophyll cycle carotenoids in Fe-deficient plants underwent epoxidations and de-epoxidations in response to ambient light conditions. In dark adapted Fe-deficient plants most of the xanthophyll cycle pigment pool was in the epoxidated form violaxanthin. We show, both by HPLC and by in vivo 505 nanometers absorbance changes, that in Fe deficient plants and in response to light, the de-epoxidated forms antheraxanthin and zeaxanthin were rapidly formed at the expense of violaxanthin. Several hours after returning to dark, the xanthophyll cycle was shifted again toward violaxanthin. The ratio of variable to maximum chlorophyll fluorescence from intact leaves was decreased by iron deficiency. However, in iron deficient leaves this ratio was little affected by light conditions which displace the xanthophyll cycle toward epoxidation or de-epoxidation. This suggests that the functioning of the xanthophyll cycle is not necessarily linked to protection against excess light input.     

Characterization of photosynthetic pigment composition, photosystem II photochemistry and thermal energy dissipation during leaf senescence of wheat plants grown in the field

Photosynthetic pigment composition and photosystem II (PSII) photochemistry were characterized during the flag leaf senescence of wheat plants grown in the field. During leaf senescence, neoxanthin and beta-carotene decreased concomitantly with chlorophyll, whereas lutein and xanthophyll cycle pigments were less affected, leading to increases in lutein/chlorophyll and xanthophyll cycle pigments/chlorophyll ratios. The chlorophyll a/b ratio also increased. With the progression of senescence, the maximal efficiency of PSII photochemistry decreased only slightly in the early morning (low light conditions), but substantially at midday (high light conditions). Actual PSII efficiency, photochemical quenching and the efficiency of excitation capture by open PSII centres decreased significantly both early in the morning and at midday and such decreases were much greater at midday than in the early morning. At the same time, non-photochemical quenching, zeaxanthin and antheraxanthin contents at the expense of violaxanthin increased both early in the morning and at midday, with a greater increase at midday. The results in the present study suggest that a down-regulation of PSII occurred in senescent leaves and that the xanthophyll cycle plays a role in the protection of PSII from photoinhibitory damage in senescent leaves by dissipating excess excitation energy, particularly when exposed to high light.     

Mechanistic aspects of the xanthophyll dynamics in higher plant thylakoids

Plant thylakoids have a highly conserved xanthophyll composition, consisting of β-carotene, lutein, neoxanthin and a pool of violaxanthin that can be converted to antheraxanthin and zeaxanthin in excess light conditions. Recent work has shown that xanthophylls undergo dynamic changes, not only in their composition but also in their distribution among Lhc proteins. Xanthophylls are released from specific binding site in the major trimeric LHCII complex of photosystem II and are subsequently bound to different sites into monomeric Lhcb proteins and dimeric Lhca proteins. In this work we review available evidence from in vivo and in vitro studies on the structural determinants that control xanthophyll exchange in Lhc proteins. We conclude that the xanthophyll exchange rate is determined by the structure of individual Lhc gene products and it is specifically controlled by the lumenal pH independently from the activation state of the violaxanthin de-epoxidase enzyme. The xanthophyll exchange induces important modifications in the organization of the antenna system of Photosystem II and, possibly of Photosystem I. Major changes consist into a modulation of the light harvesting efficiency and an increase of the protection from lipid peroxidation. The xanthophyll cycle thus appears to be a signal transduction system for co-ordinated regulation of the photoprotection mechanisms under persistent stress from excess light.     

Accumulation of macular xanthophylls in unsaturated membrane domains

The distribution of macular xanthophylls, lutein and zeaxanthin, between domains formed in membranes made from an equimolar ternary mixture of dioleoylphosphatidylcholine/sphingomyelin/cholesterol, called a raft-forming mixture, was investigated. In these membranes, two domains are formed: the raft domain enriched in saturated lipids and cholesterol (detergent-resistant membranes, DRM), and the bulk domain enriched in unsaturated lipids (detergent-soluble membranes, DSM). These membrane domains have been separated using cold Triton X-100 extraction from membranes containing 1 mol% of either lutein or zeaxanthin. The results indicated that xanthophylls are substantially excluded from DRM and remain concentrated in DSM. Concentrations of xanthophylls in DRM and DSM calculated as the mole ratio of either xanthophyll to phospholipid were 0.005 and 0.03, respectively, and calculated as the mole ratio of either xanthophyll to total lipid (phospholipid + cholesterol) were 0.003 and 0.025, respectively. Thus, xanthophylls are over eight times more concentrated in DSM than in DRM. No significant difference in the distribution of lutein and zeaxanthin was found. It was also demonstrated using saturation-recovery EPR that at 1 mol%, neither lutein nor zeaxanthin affect the formation of membrane domains. The location of xanthophylls in domains formed from unsaturated lipids is ideal if they are to act as a lipid antioxidant, which is the most accepted mechanism through which lutein and zeaxanthin protect the retina from age-related macular diseases.     

The xanthophyll cycle in green algae (chlorophyta): its role in the photosynthetic apparatus

Light-dependent conversion of violaxanthin to zeaxanthin, the so-called xanthophyll cycle, was shown to serve as a major, short-term light acclimation mechanism in higher plants. The role of xanthophylls in thermal dissipation of surplus excitation energy was deduced from the linear relationship between zeaxanthin formation and the magnitude of non-photochemical quenching. Unlike in higher plants, the role of the xanthophyll cycle in green algae (Chlorophyta) is ambiguous, since its contribution to energy dissipation can significantly vary among species. Here, we have studied the role of the xanthophyll cycle in the adaptation of several species of green algae (Chlorella, Scenedesmus, Haematococcus, Chlorococcum, Spongiochloris) to high irradiance. The xanthophyll cycle has been found functional in all tested organisms; however its contribution to non-photochemical quenching is not as significant as in higher plants. This conclusion is supported by three facts: (i) in green algae the content of zeaxanthin normalized per chlorophyll was significantly lower than that reported from higher plants, (ii) antheraxanthin + zeaxanthin content displayed different diel kinetics from NPQ and (iii) in green algae there was no such linear relationship between NPQ and Ax + Zx, as found in higher plants. We assume that microalgae rely on other dissipation mechanism(s), which operate along with xanthophyll cycle-dependent quenching.     

The current concepts of functional role of carotenoids in the eukaryotic chloroplasts

Current state of knowledge of functional role of carotenoids in algal and higher plant chloroplasts is reviewed. Basic functions of carotenoids are shown to be light-protective, light-absorbing, and structural, as well as participating in photochemical processes of photosystems I and II. Such xanthophylls as neoxanthin, fucoxanthin, peridinin and alloxanthin, which have allenic or acetylenic bond, mostly function as light-absorbers. They transmit absorbed energy to chlorophyll b. Other xanthophylls occupying certain strictly specified loci in light-absorbing chlorophyll-a/b-protein complexes of photosystems have either structural function (lutein) or light-protective function (zeaxanthin, antheraxanthin, violaxanthin). Carotenoids of xanthophyll cycles preserve chlorophylls and lipids of photosynthetic membranes from photodestruction at overlighting in the presence of oxygen. In eukaryotic chloroplasts, three types of xanthophyll cycles were found: violaxanthin, lutein-5,6-epoxide, and diadinoxanthin. The similarities and dissimilarities between epoxidation and de-epoxidation reactions of these cycles are discussed in detail in the present work. The pattern of occurrence of xanthophyll cycles among higher plants and freshwater and marine algae is outlined.     

The P450-type carotene hydroxylase PuCHY1 from Porphyra suggests the evolution of carotenoid metabolism in red algae

Carotene hydroxylases catalyze the hydroxylation of a-and b-carotene hydrocarbons into xanthophylls. In red algae, b-carotene is a ubiquitously distributed carotenoid, and hydroxylated carotenoids such...     

Xanthophyll cycle pool size and composition in relation to the nitrogen content of apple leaves

The objective of this study was to determine xanthophyll cycle pool size and composition in response to N status and their relationships to non-photochemical quenching in apple leaves. Bench-grafted Fuji/M.26 trees were fertilized with different N concentrations (0-20 mM) in a modified Hoagland's solution for 6 weeks to create a wide range of leaf N status (1-4.4 g m(-2)). Chlorophyll content, xanthophyll cycle pool size, lutein, total carotene, and neoxanthin on a leaf area basis all increased linearly with increasing leaf N. However, only the ratios of the xanthophyll cycle pool and of lutein to chlorophyll were higher in low N leaves than in high N leaves. Under high light at midday, both zeaxanthin (Z), expressed on a chlorophyll basis, and the percentage of the xanthophyll cycle pool present as Z, increased as leaf N decreased. Thermal dissipation of excitation energy, measured as non-photochemical quenching of chlorophyll fluorescence, was positively related to, whereas efficiency of excitation transfer and photosystem II quantum efficiency were negatively related to, Z, expressed on a chlorophyll basis or on a xanthophyll cycle pool basis. It is concluded that both xanthophyll cycle pool size (on a chlorophyll basis) and conversion of violaxanthin to zeaxanthin are enhanced in response to N limitation to dissipate excessive absorbed light under high irradiance.     

Genetic manipulation of carotenoid biosynthesis and photoprotection

There are multiple complementary and redundant mechanisms to provide protection against photo-oxidative damage, including non-photochemical quenching (NPQ). NPQ dissipates excess excitation energy as heat by using xanthophylls in combination with changes to the light-harvesting complex (LHC) antenna. The xanthophylls are oxygenated carotenoids that in addition to contributing to NPQ can quench singlet or triplet chlorophyll and are necessary for the assembly and stability of the antenna. We have genetically manipulated the expression of the epsilon-cyclase and beta-carotene hydroxylase carotenoid biosynthetic enzymes in Arabidopsis thaliana. The epsilon-cyclase overexpression confirmed that lut2 (lutein deficient) is a mutation in the epsilon-cyclase gene and demonstrated that lutein content can be altered at the level of mRNA abundance with levels ranging from 0 to 180% of wild-type. Also, it is clear that lutein affects the induction and extent of NPQ. The deleterious effects of lutein deficiency on NPQ in Arabidopsis and Chlamydomonas are additive, no matter what the genetic background, whether npq1 (zeaxanthin deficient), aba1 or antisense beta-hydroxylase (xanthophyll cycle pool decreased). Additionally, increasing lutein content causes a marginal, but significant, increase in the rate of induction of NPQ despite a reduction in the xanthophyll cycle pool size.     

Variations in the carotenoid content of Chlamydomonas reinhardii throughout the cell cycle

Synchronous cultures of Chlamydomonas reinhardii have been examined for the total amounts of carotenoid and chlorophyll present throughout a 12 hrs light–4 hrs dark life cycle. Variations in the carotenoid distribution at different points within the cell cycle have been found. During the greater part of the light period all major carotenoids increased at a proportionally similar rate. However, the increases in lutein and violaxanthin preceded those in β-carotene and neoxanthin by some 2 hrs and that in loroxanthin, an algal xanthophyll, by about 3 hrs. A marked drop in total carotenoid accumulation, corresponding to similar temporary falling away in the accumulation of β-carotene, lutein and violaxanthin occurred at 9 hrs. The correspondence of this with the established drop in RNA accumulation and the break-up of the nucleolus was pointed out. Considerable redistribution among the carotenoids occurred during the dark period, notably the amount of β-carotene increased relative to the total xanthophylls. The full significance of these results can not be estimated in the absence of comparative data on related organisms.     

The binding of Xanthophylls to the bulk light-harvesting complex of photosystem II of higher plants. A specific requirement for carotenoids with a 3-hydroxy-beta-end group

The pigment composition of the light-harvesting complexes (LHCs) of higher plants is highly conserved. The bulk complex (LHCIIb) binds three xanthophyll molecules in combination with chlorophyll (Chl) a and b. The structural requirements for binding xanthophylls to LHCIIb have been examined using an in vitro reconstitution procedure. Reassembly of the monomeric recombinant LHCIIb was performed using a wide range of native and nonnative xanthophylls, and a specific requirement for the presence of a hydroxy group at C-3 on a single beta-end group was identified. The presence of additional substituents (e.g. at C-4) did not interfere with xanthophyll binding, but they could not, on their own, support reassembly. cis isomers of zeaxanthin, violaxanthin, and lutein were not bound, whereas all-trans-neoxanthin and different chiral forms of lutein and zeaxanthin were incorporated into the complex. The C-3 and C-3' diols lactucaxanthin (a carotenoid native to many plant LHCs) and eschscholtzxanthin (a retro-carotenoid) both behaved very differently from lutein and zeaxanthin in that they would not support complex reassembly when used alone. Lactucaxanthin could, however, be bound when lutein was also present, and it showed a high affinity for xanthophyll binding site N1. In the presence of lutein, lactucaxanthin was readily bound to at least one lutein-binding site, suggesting that the ability to bind to the complex and initiate protein folding may be dependent on different structural features of the carotenoid molecule. The importance of carotenoid end group structure and ring-to-chain conformation around the C-6-C-7 torsion angle of the carotenoid molecule in binding and complex reassembly is discussed.     

Iron deficiency-induced changes in the photosynthetic pigment composition of field-grown pear (Pyrus communis L) leaves

In this work we characterize the changes induced by iron deficiency in the pigment composition of pear (Pyrus communis L.) leaves grown under high light intensities in field conditions in Spain. Iron deficiency induced decreases in neoxanthin and β-carotene concomitantly with decreases in chlorophyll a, whereas lutein and carotenoids within the xanthophyll cycle were less affected. Iron deficiency caused major increases in the lutein/chlorophyll a and xanthophyll cycle pigments/chlorophyll a molar ratios. The chlorophyll a/chlorophyll b ratio increased in response to iron deficiency. The carotenoids within the xanthophyll cycle in iron-deficient and in iron-sufficient (control) leaves underwent epoxidations and de-epoxidations in response to ambient light conditions. In control leaves dark-adapted for several hours, most of the xanthophyll cycle pigment pool was in the epoxidated form vio-laxanthin, whereas iron-deficient leaves had significant amounts of zeaxanthin. Iron-deficient leaves also exhibited an increased non-photochemical quenching, supporting the possibility of a role for pigments within the xanthophyll cycle in photoprotection.