Photosystem reaction centers and structural organization of chloroplasts in chlorotica mutants of Pisum sativum L

Chlorophyll-protein complexes of photosystems (PS), as well as ultrastructural arrangement of chloroplasts in pea leaves of the primary cultivar Torsdag and mutants chlorotica 2004 and 2014 were studied. It was shown that both mutants accumulated 80 and 55% of chlorophyll, respectively, and were able to synthesize all of four types of photosystems complexes. The value of the light-harvesting antenna in mutant 2014 was close to the control one, and in mutant 2004 it increased significantly (by 30%). These changes were caused by a proportional decrease, 40-50%, of any complexes in mutant 2014, whereas the number of PS I reaction centers in mutant 2004 decreased by 50% and the reaction centers of PS II complexes were almost completely retained. It was established that the proportional decrease of PS I and PS II complexes in mutant chlorotica 2014 was followed by a partial reduction of the entire membrane system in chloroplasts, but with a good development of both granal and intergranal sites of thylakoids. On the contrary, the loss of complexes of PS I reaction centers in mutant chlorotica 2004 led to a reduction of unstacked sites of thylakoids in chloroplasts. It was concluded that the disturbance of the lateral orientation of the membrane system of chloroplasts is associated with the loss of complexes of reaction centers of PS I, which is predominantly localized in unstacked sites of thylakoids.     

The Involvement of the Complexes of Photosystems in the Organization of Chloroplast Ultrastructure in Pea Mutants

We studied the involvement of pigment-protein complexes of photosystems (PS) in the development and spatial arrangement of thylakoids in chloroplasts of pea (Pisum sativum L.) leaves. The initial line (cv. Torsdag) and its mutants, chlorotica 2004 displaying primary disturbances in the PSI reaction centers and chlorotica 2014 containing only 50% of chlorophyll and, as a sequence, the reduced amount of all pigment-protein complexes. A proportional decrease in the content of PSI and PSII complexes in the chlorotica 2014 mutant resulted in a partial reduction of the whole chloroplast membrane system, whereas grana and stroma thylakoid regions were well developed. In contrast, a loss of only 20% of chlorophyll and destruction of PSI complexes in the chlorotica 2004 mutant by 50% resulted in the destruction of stroma thylakoid regions and disturbed longitudinal thylakoid and grana orientation. It was concluded that protein-protein interactions in pigment-protein complexes played a key role in the structure of thylakoid membranes and their longitudinal orientation.     

Photosystem Damage and Spatial Architecture of Thylakoids in Chloroplasts of Pea Chlorophyll Mutants

The effect of chlorophyll–protein complexes on the ultrastructure of chloroplasts was studied in the leaves of pea, the parent cultivar Torsdag and mutants chlorotica 2004 and 2014. The mutants were shown to accumulate 80 and 55% of chlorophyll, relative to the control, while the composition of the synthesized photosystem complexes was the same as in the parent cultivar Torsdag. The size of the light-harvesting antenna was similar to the control in the 2014 mutant but considerably increased (by 30%) in the 2004 mutant. These changes were due to a proportional decrease in the number of all complexes (by 40–45%) in the 2014 mutant. At the same time, the number of reaction center complexes of photosystem I (PS I) decreased by 50% while that of photosystem II (PS II) remained virtually constant in the 2004 mutant. A proportional decrease in the number of the PS I and PS II complexes in the chlorotica 2014 mutant was accompanied by a partial reduction of the entire chloroplast membrane system against the background of normal development of both granal and intergranal sites of thylakoids. Conversely, the loss of PS I reaction centers led mainly to the reduction of the intergranal sites of thylakoids in chloroplasts. This effect is attributed to the prevalence of PS I complexes in the intergranal thylakoids.     

Structural-functional organization of chloroplasts in leaves of xantha-702 mutant of Gossypium hirsutum L

For cotton mutant xantha (Gossypium hirsutum L.), it has been established that synthesis of 5-aminolevulinic acid was blocked in the light. In the light this mutant accumulates chlorophyll by 30 times lower as compared to the parent type. In mutant xantha, a very few pigment-protein complexes of PS-I and PS-II are formed in chloroplasts, and formation of membrane system in these is blocked at the early stages, in most cases, at the stage of bubbles and single short thylakoids. Functional activity of reaction centers of PS-I and PS-II is close to zero. Only light-harvesting chlorophyll-a/b protein complexes of the two photosystems are formed in mutant xantha plastid membranes with maximum chlorophyll fluorescence at 728 and 681 nm, respectively. It has been concluded that in mutant xantha genetic block of 5-aminolevulinic acid biosynthesis in the light disturbs the formation and functioning of the complexes of reaction centers of PS-I and PS-II, hindering the development of the whole membrane system in chloroplasts, causing a sharp decrease in productivity.     

Spectral characteristics and the structure of chloroplasts upon blocking the early stages of chlorophyll biosynthesis

The cotton mutant xantha (Gossypium hirsutum L.) with the blocked synthesis of 5-aminolevulinic acid in the light has been shown to accumulate chlorophyll 30 times less than the parent type. In chloroplasts of the mutant xantha, the formation of the membrane system is blocked at the earliest stages, mainly at the stage of bubbles and single short thylakoids. Only light-harvesting chlorophyll-a/b-protein complexes I and II with chlorophyll fluorescence maxima at 728 and 681 nm, respectively, are formed in plastid membranes of the mutant. It has been concluded that the genetic block of chlorophyll biosynthesis in the mutant xantha disturbs the formation and functioning of the complexes in reaction centers of PS-I and PS-II, inhibiting the development of the whole membrane system of chloroplasts at the stage of bubbles and single thylakoids.     

Fluorescence,chlorophyll shape and number of photosystem reaction centers I and II in chlorotica mutants of Pisum sativum L

The fluorescent and absorbing properties of chloroplasts and pigment-protein complexes isolated by gel electrophoresis from pea leaves of the cultivar Torsdag and the mutants chlorotica 2004 and 2014 were studied. From the absorption and fluorescence spectra of chlorophylls and their 2nd derivatives, the range of their changes in the native state at 23 degrees C and specific maxima of fluorescence and the forms of chlorophyll of individual complexes at -196 degrees C were found. It was found that in mutant chlorotica 2004 the intensity of fluorescence of long-wave band at 745 nm (23 degrees C) and the maximum--at 728 nm (-196 degrees C) belonging to the light-harvesting complex I increased. Nevertheless, the accumulation of the chlorophyll forms in this mutant at 690, 697 and 708 nm, which make an antenna of reaction centers of photosystem (PS) I decreased. No spectral differences from the spectrum of the wild type were found in mutant chlorotica 2014, except for a weakening of interaction between the complexes of PS I and PS II. It was shown by gel electrophoresis that both mutants were capable of synthesizing any chlorophyll-protein complexes. However, the analysis of the photochemical activity of reaction centers of PS I and PS II as well as calculations of the value of the photosynthetic unit and the number of reaction centers of the photosystems enabled us to conclude that the quantity of the reaction centers of PS I in the mutant chlorotica 2004 was 1.7 times lower due to disturbance of mutations in biosynthesis or the formation of the chlorophyll a-protein complex of PS I. No primary effect of mutation of chlorotica 2014 was established. Proportional changes of all parameters in this mutant gave us the ground to consider them as secondary ones, which are caused by a decrease in chlorophyll content by half.     

Spectral properties and the number of photosynthetic reaction centers in chlorophyll-deficient mutants of <Emphasis Type="Italic">Pisum sativum</Emphasis>

Spectral and photochemical properties were analyzed on intact chloroplasts and pigment-protein complexes isolated with gel electrophoresis from pea (Pisum sativum L.) leaves of parental variety Torsdag and of chlorophyll-deficient mutants chlorotica 2004 and 2014. Measurements of chlorophyll absorption and fluorescence spectra and of second derivative low-temperature (–196°C) spectra clarified exact positions of fluorescence maxima and revealed the chlorophyll forms of individual complexes in samples investigated. The chlorotica 2004 mutant, whose hybrids yield the heterosis effect, was characterized by the decreased accumulation of chlorophyll forms absorbing at 690, 697, and 708 nm, known to constitute the core antenna in the vicinity of photosystem I (PSI) reaction center. In the chlorotica 2014 mutant, whose hybrids are low productive, the interaction between PSI and PSII complexes was weakened, but no other difference from the parental variety was observed. The analysis of PSI and PSII photochemical activities, as well as estimates of light-harvesting antenna size and the number of reaction centers revealed that the chlorotica 2004 mutant is deficient in the number of PSI reaction centers by a factor of 1.7. This deficiency resulted from the mutation-induced disorder in biosynthesis of chlorophyll a-protein complex of PSI. It appears that gene interactions between the 2004 mutant and the parental variety Torsdag enhance the functional and metabolic activity of leaves in their hybrids, thereby yielding the heterosis effect.Translated from Fiziologiya Rastenii, Vol. 52, No. 2, 2005, pp. 172–183.Original Russian Text Copyright © 2005 by Ladygin, Vaishlya.This revised version was published online in April 2005 with a corrected cover date.     

Pigment accumulation and functional activity of chloroplasts in common Pisum sativum L. mutants with low chlorophyll level (chlorotica)

Pea mutants chlorotica 2004 and 2014 with a low content of chlorophyll were studied. The mutant 2004 has light green leaves and stem, and the mutant 2014 has yellow green leaves and stem. They accumulate approximately 80 and 50% chlorophylls of the parent form of pea Torsdag cv. The content of carotene in carotenoids of the mutant 2004 was much lower, and the accumulation of lutein and violaxanthine was increased. The accumulation of all carotenoids in the mutant 2014 decreased almost proportionally to a decrease in the chlorophyll content. The rate of CO2 evolution in mutant chlorotica 2004 and 2014 was established to be lower. The quantum efficiency of photosynthesis in the mutants was 29-30% lower as compared to the control, and in hybrid plants it was 1.5-2-fold higher. It is assumed that the increase in the activity of the night-time respiration in gas exchange of chlorotica mutants and the drop of photosynthesis lead to a decrease in biomass increment. The results obtained allow us to conclude that the mutation of chlorotica 2004 and 2014 affects the genes controlling the formation and functioning of different components of the photosynthetic apparatus.     

Optimization of isolation conditions for three types of pigment protein-lipid complexes from chloroplasts during solubilization with Triton X-100]

The maximal total release of pigment protein-lipid complexes (PPLC) during their isolation from pea chloroplasts was achieved by 1-hr solubilization with Triton X-100, the Triton:chlorophyll (T/Chl) ratio being 50 mg/mg/ml. The total yield of the reaction center complexes (sigma PPLC RC) was 22,3%, whereas that of the auxiliary light-accumulating complex (ALA-PPLC) was approximately 32% with respect to Chl. An increase in the solubilization time and of the T/Chl ratio resulted in dissociation of ALA-PPLC. On the contrary, the reaction center complexes steadily maintained their composition and high photochemical activity within a wide range of T/Chl during 24--28 hrs of solubilization. The purest preparations of PPLC RC of phostosystem I (PS-I) were obtained by 24 hr-incubation (T/Chl = 80); their Chl/P700 ratio after a single fractionation on DEAE-cellulose was equal to 36. A considerable increase of T/Chl and of the solubilization time hampered the chromatographical separation of PPLC RC of PS-I and PPLC RC of PS-II. The optimal conditions for isolation of PPLC RC of PS-I and PPLC RC of PS-II were: solubilization at T/Chl 80--120 and prolongation of incubation time from 5 to 7 hrs. The photochemical activity of the complexes obtained was maximal and correlated with the minimal content of admixture P700 (1 molecule of P700 per 450--500 molecules of Chl.).     

Functional effects of chemical modification of unstacked and stacked chloroplasts with glutaraldehyde.

Although glutaraldehyde alkylates protein NH₂ groups to the same extent in unstacked and stacked thylakoids, the photosynthetic electron transport of the stacked membranes is always more inhibited. Inhibition of photosystem II electron transport, measured in the presence of lipophilic Hill oxidants, is 20–30% in unstacked and 60–70% in stacked thylakoids. Photosystem I electron transport is nearly completely inhibited in both preparations, but in the case of stacked thylakoids maximal inhibition occurs at a lower glutaraldehyde level than in unstacked thylakoids. In contrast, the photooxidation of the reaction center chromophore of photosystem I (P700) is unaffected by the glutaraldehyde treatment of either stacked or unstacked chloroplasts. The results are discussed with regard to the accessibility of membrane sites to exogenous electron transport cofactors, in view of the observation that N-methylphenazonium methosulfate, a quencher of electronically excited chlorophyll a, partitions more easily into the pigment domains of the glutaraldehyde-fixed unstacked thylakoids.     

Pigment–Protein Complexes and the Number of Reaction Centers of the Photosystems in Pea Chlorophyll Mutants

We studied fluorescent and absorption properties of the chloroplasts and pigment–protein complexes isolated by gel electrophoresis from the leaves of pea, the parent cultivar Torsdag and mutants chlorotica 2004 and 2014. Specific fluorescence peaks of chlorophyll forms in individual complexes have been determined from the absorption and fluorescence spectra of the chloroplast chlorophyll and their second derivatives at 23 and –196°C. The mutant chlorotica 2004 proved to have an increased intensity of a long-wave band of the light-harvesting complex I at both 23°C (745 nm) and –196°C (728 nm). At the same time, this mutant manifested a decreased accumulation of the chlorophyll forms making up the nearest-neighbor antenna of the PS I reaction center (at 690, 697, and 708 nm). No spectral differences have been revealed between chlorotica 2014 mutant and the parent cultivar. Gel electrophoresis revealed the synthesis of all chlorophyll–protein complexes in both mutants. At the same time, analysis of photochemical activity of PS I and PS II reaction centers and calculations of their number and the size of the light-harvesting antenna have shown that the number of reaction centers in the PS I of chlorotica 2004 mutant is reduced by a factor of 1.7 because its chlorophyll a–protein complex is disturbed by the mutation. The primary effect of chlorotica 2014 mutation remains unclear. The proportional changes in the content of photosystem complexes in this mutant suggest that they are secondary and result from a 50% decrease in chlorophyll content.     

Disturbed structure and function of chloroplasts during blocked biosynthesis of 5-aminolevulinic acid in the light

Xantha-702 mutant of cotton (Gossypium hirsutum L.) proved to have blocked synthesis of 5-aminolevulinic acid in the light. Accordingly, mutant leaves accumulated 2–5% chlorophyll of baseline. Mutant plants demonstrated disturbed production of pigment-protein complexes of photosystems I (PSI) and II (PSII) and generation of the chloroplast membrane system blocked at the early stages, largely, at the stages of vesicles and single short thylakoid. The functional activity of the PSI and PSII reaction centers was close to zero. Only the chlorophyll a/b light-harvesting complexes of PSI and PSII with the chlorophyll fluorescence peaks at 728 and 681 nm, respectively, were produced in the xantha-702 mutant. We propose that the genetic block of 5-aminolevunilic acid biosynthesis in the light in the xantha-702 mutant disturbs the formation and activity of the complexes of the reaction centers of PS-I and PS-II and inhibits the development of the whole membrane system of chloroplasts.     

Action spectra and functional antenna sizes of Photosystems I and II in relation to the thylakoid membrane organization and pigment composition

The functional organization of competent photosynthetic units in developing thylakoids from intermittent-light grown pea as well as in the unstacked, stacked and phosphorylated stacked thylakoids from its mature chloroplasts was characterized by polarographic measurements of action spectra, reaction centre contents and optical cross-sections for PS I-mediated O2 uptake and PS II-mediated O2 evolution. The minimum antenna sizes of 60 and 37 chlorophyll a molecules for PS I and PS II, respectively, were determined in developing thylakoids with a ratio of Chl a/Chl b>50. In mature chloroplasts, the embedded light-harvesting chlorophyll a/b-binding (LHC) protein complexes increased the PS I and PS II effective antenna sizes by 3–6 times depending on the thylakoid membrane organization. In unstacked thylakoids, a randomization of PS I, PS II and LHC II led to the most uniform spectral distribution of light harvesting between the two photosystems but caused the maximal difference of their antenna sizes to be 370 and 100 Chls for the competent PS I and PS II units, respectively. Following the Mg2+-induced stacking of thylakoids, opposite complementary changes of the action spectra, antenna sizes and Chl a/Chl b ratios indicated a redistribution of a LHC II pool of 100 Chl ( a + b) molecules from PS I to PS II. Unlike to the stroma-exposed PS II in unstacked thylakoids, the granal PS II units of 200 Chls demonstrated an additional 2-fold increase of the effective antenna size due to energy transfer within PS II dimers under strong background illumination, which closed >90% of reaction centres. Protein phosphorylation of the stacked thylakoids induced a significant inactivation of the O2-evolving PS II centres but did not cause complementary changes of the action spectra and antenna sizes of the competent PS I and PS II. In this case, light harvesting parameters of the O2-evolving PS II units were nearly unaffected, whereas the obvious relative increase of the PS I activity at 650 nm and its decrease at >700 nm both in the action spectrum and optical cross-section measurements might suggest a substitution of PS I units in the O2-reducing fraction by another distinct fraction of -type which in turn is not the same to PS I units in unstacked thylakoids.     

Cold-acclimation protects photosystem II against freezing damage in the halotolerant alga Dunaliella salina

Cold-acclimation (CA) of the halotolerant alga Dunaliella was inhibited by light and by high salt. CA was associated with enhanced resistance to freezing in saline growth solutions, as manifested by protection of photosynthetic oxygen evolution and by reduced permeabilisation of the plasma membrane. Oxygen evolution activity in isolated chloroplasts was not affected by freezing, but was inhibited by high salt and the inhibition could be reversed or protected by glycerol. The activity of chloroplasts from cold-acclimated cells was more resistant to salt than of non-acclimated cells. Electron transport measurements in chloroplasts indicated that high salt inhibited PS-II, but not PS-I electron transport. High salt also inhibited PS-II thermoluminescence (TL) activity in chloroplasts. Similar inhibition of PS-II TL was observed by freezing intact cells in saline solutions. Chloroplasts from cold-acclimated cells had enhanced resistance to inhibition of PS-II electron transport and of PS-II TL by high salt. These results suggest that inhibition of oxygen evolution upon freezing Dunaliella cells may result from inactivation of PS-II due to massive influx of salt and loss of glycerol. The enhanced freeze-resistance of cold-acclimated cells to inhibition of oxygen evolution can be accounted for partly by protection of PS-II against high salt.     

Oxygen-evolving photosystem II preparation from wild type and photosystem II mutants of Synechocystis sp. PCC 6803.

We present here a simple and rapid method which allows relatively large quantities of oxygen-evolving photosystem II- (PS-II-) enriched particles to be obtained from wild-type and mutants of the cyanobacterium Synechocystis 6803. This method is based on that of Burnap et al. [Burnap, R., Koike, H., Sotiropoulou, G., Sherman, L. A., & Inoue, Y. (1989) Photosynth. Res. 22, 123-130] but is modified so that the whole preparation, from cells to PS-II particles, is achieved in 10 h and involves only one purification step. The purified preparation exhibits a 5-6-fold increase of O2-evolution activity on a chlorophyll basis over the thylakoids. The ratio of PS-I to PS-II is about 0.14:1 in the preparation. The secondary quinone electron acceptor, QB, is present in this preparation as demonstrated by thermoluminescence studies. These PS-II particles are well-suited to spectroscopic studies as demonstrated by the range of EPR signals arising from components of PS-II that are easily detectable. Among the EPR signals presented are those from a formal S3-state, attributed to an oxidized amino acid interacting magnetically with the Mn complex in Ca(2+)-deficient PS-II particles, and from S2 modified by the replacement of Ca2+ by Sr2+. Neither of these signals has been previously reported in cyanobacteria. Their detection under these conditions indicates a similar lesion caused by Ca2+ depletion in both plants and cyanobacteria. The protocol has also been applied to mutants which have site-specific changes in PS-II. Data are presented on mutants having changes on the electron donor (Y160F) and electron acceptor (G215W) side of the D2 polypeptide.     

Immunogold localization of photosystem I and photosystem II light-harvesting complexes in cryptomonad thylakoids

Summary— The molecular organization of the thylakoids of Cryptomonas rufescens was studied by immunoelectron microscopy employing antibodies against photosystem (PS)-I and two PS-II antenna proteins. The PS-I complex and the 19-kDa chlorophyll a/c light-harvesting (LH) protein are both localized along the length of the thylakoid membranes. The external membranes of the paired thylakoids are enriched in PS-I whereas the chlorophyll a/c LH protein is more concentrated in the internal or appressed membranes. However, unlike the situation in higher plants and Chlamydomonas, there is not a marked asymmetry in the concentration of PS-I and chorophyll a/c LH protein in the two types of membranes. Double labelling studies of sections and isolated PE-PS-II particles with anti-phycoerythrin and anti-LH confirmed that phycoerythrin is localized in the thylakoid lumen and that this pigment exists in two forms, a fraction closely associated with the thylakoid membranes and another fraction free in the lumen. These results confirm the uniqueness of cryptomonad thylakoids.     

Evolution of photosynthetic reaction centers

The common ancestor of all photosynthetic prokaryotes and organelles contained chlorophyll (Chl) a. All green and purple photosynthetic bacteria descended from a common bacteriochlorophyll (Bchl) a-containing ancestor which diverged from the Chl a line. Separate PS-I and PS-II reaction centers may have evolved before the appearance of Bchl a. When the transition to Bchl a occurred, the resultant organism contained two types of reaction center, “PS-I” and “PS-II.” One line of development eliminated “PS-II” and evolved into the green bacteria. The other line eliminated “PS-I” and became the purple bacteria. In the Chl a-containing organisms the evolution of PS-II continued until oxygen evolution was achieved.     

A test of the binary chloroplast membrane hypothesis by using a nonpenetrating chemical probe,p-(diazonium)-benzenesulfonic acid

Summary Spinach chloroplasts were exposed to³⁵S-labeledp-(diazonium)-benzenesulfonic acid (DABS), a water soluble compound which does not penetrate lipophilie regions of membranes, and which is highly reactive toward amino acid functionagroups such as -amino, sulfhydryl, histidine, and tyrosine groups. Amino groups inl lipids can also form similar, stable covalent bonds by diazo coupling. Both chloroplast lipids and proteins were labeled with DABS, the total binding being about 1 DABS per 10 chlorophylls, depending on the reaction conditions.After diazo coupling and subsequent digitonin fractionation into photosystems I and II enriched fractions, it was observed that PS-I was more highly labeled than PS-III usually by a factor of 10 to 24 times (on a per chlorophyll basis). After digitonin isolation, however, the PS-II portion bound an amount of DABS similar to the PS-I binding, We interpret these data as consistent with the binary membrane hypothesis (Arntzen. Dilley and Crane (1969),J. Cell Biol. 43:16), which visualizes PS-I on the externa, half of a 90 Å grana membrane, and PS-II occurring on the interior half of thel membrane. The alternative explanation that PS-II and PS-I are arranged as a mosaic, and that the low DABS binding in PS-II is caused by burial of the diazo reactive groups in the interior of the proteins (and only exposed through the denaturing effect of digitonin) is not directly ruled out. However, this alternative is not consistent with the facts that: (a) most of the membrane proteins in PS-I and PS-II are identical in electrophoretic properties and therefore probably have similar overall structures; and (b) digitonin does not lead to appreciable denaturation of proteins, evidenced by the retention of PS-II electron transport activity.     

Correlation of the photosystem I and II reaction center chlorophyll-protein complexes, CP-a1 and CP-aII with photosystem activity and low temperature fluorescence emission properties in mutants of Scenedesmus

Abstract Comparative studies on the low temperature fluorescence emission of whole cells, purified chlorophyll-protein (CP) complexes and on patterns noted in sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) for chlorophyll-protein complexes and chloroplast membrane polypeptides of Scenedesmus obliquus with several distinct mutant classes has allowed further correlation between the fluorescence emission bands seen at 77K and the known chlorophyll-protein complexes. In mutants deficient in photosystem II (PS-II; total loss of the reducing side) the fluorescence emission spectra showed only two peaks, i.e., 686 and 718 nm, but in the wild type, in mutants lacking the oxidizing side of PS-II and in phenotypes missing the CP-a₁ complex (and P-700 activity) all three emission bands at 686, 696 and 718 nm were present. In a mutant lacking the light-harvesting CP-a/b complex the emission peak at 686 nm was strongly reduced and the longer wavelength emissions predominated. Gel electrophoresis studies showed that the PS-II (reducing side) mutants lacked the polypeptides of apparent molecular weight 54 and 51 kilodaltons and the chlorophyll-protein complex, CP-a_II, of apparent molecular weight 32 kilodaltons. Contrarily, the loss of the oxidizing side of PS-II did not result in any alteration of these components. Genetic deletion of CP-a₁ did not alter significantly the long wavelength emission even though the isolated CP-a₁ shows the low temperature-dependent long wavelength emission comparable to that seen in the whole cell. It was deduced that remaining PS-I antennae chlorophylls must account for the emission seen at 718 nm. The absence of the CP-a/b complex and the strong simultaneous decrease of the 686 nm emission strengthens the concept that this complex is the primary emitter of fluorescence at room temperature. Its absence facilitated the detection of the CP-a_II complex in SDS-PAGE and enhanced the in vivo fluorescence by the two photosystems. Parallel experiments with two mutants which green and develop, one to the wild-type and the other to the CP-a/b deficient phenotype, provided additional evidence for the source of the low temperature emission bands.     

Glucans from alkaline extract of a hybrid mushroom (backcross mating between PfloVv12 and Volvariella volvacea): structural characterization and study of immunoenhancing and antioxidant properties

Two different glucans (water-soluble PS-I, water-insoluble PS-II) were isolated from the alkaline extract of the fruit bodies of hybrid mushroom. PS-I was found to consist of only (1→6)-linked β-D-glucopyranose. PS-II was composed of terminal, (1→3,4)-linked, and (1→3)-linked β-D-glucopyranosyl moieties in a molar ratio of nearly 1:1:1. PS-I showed macrophages, splenocytes, and thymocytes activation as well as antioxidant property. On the basis of sugar analysis, methylation analysis, and NMR studies ((1)H, (13)C, DEPT-135, TOCSY, DQF-COSY, NOESY, ROESY, HMQC, and HMBC), the structure of the repeating unit of these glucans were established as: