The effects of external NaCl on thylakoid stacking in lettuce plants

The average degree of thylakoid stacking was determined for loose-leaf lettuce plants which were grown in complete nutrient solutions containing either 10 or 100mol m^?3 NaCl. Digitonin fractionation and differential centrifugation were used to assay the level of thylakoid stacking. Based on a comparison between 10mol m^?3 NaCl-grown and 100mol m^?3 NaCl-grown lettuce plants of equal ages, digitonin assays indicated that significantly less stacking occurred in 100mol m^?3 NaCl-grown plants. Isolated thylakoid membranes from 100mol m^?3 NaCl-grown plants were also characterized by a greater capacity to absorb divalent cations and by a higher chlorophyll a/b ratio. Since plants from both growth salinities were capable of a marked increase in thylakoid stacking upon a transition from high to low irradiance, the observed differences in thylakoid stacking were not attributed to a salinity-related impairment of stacking mechanisms. Instead, the salinity-induced differences in thylakoid stacking appear to represent a process of controlled adjustment.     

Pigment composition and functional state of the thylakoid membranes during preparation of samples for pigment-protein complexes separation by nondenaturing gel electrophoresis

The present study was conducted to examine changes in photosynthetic pigment composition and functional state of the thylakoid membranes during the individual steps of preparation of samples that are intended for a separation of pigmentprotein complexes by nondenaturing polyacrylamide gel electrophoresis. The thylakoid membranes were isolated from barley leaves (Hordeum vulgare L.) grown under low irradiance (50 μmol m⁻² s⁻¹). Functional state of the thylakoid membrane preparations was evaluated by determination of the maximal photochemical efficiency of photosystem (PS) II (F_V/F_M) and by analysis of excitation and emission spectra of chlorophyll a (Chl a) fluorescence at 77 K. All measurements were done at three phases of preparation of the samples: (1) in the suspensions of osmotically-shocked broken chloroplasts, (2) thylakoid membranes in extraction buffer containing Tris, glycine, and glycerol and (3) thylakoid membranes solubilized with a detergent decyl-β-D-maltosid. F_V/F_M was reduced from 0.815 in the first step to 0.723 in the second step and to values close to zero in solubilized membranes. Pigment composition was not pronouncedly changed during preparation of the thylakoid membrane samples. Isolation of thylakoid membranes affected the efficiency of excitation energy transfer within PSII complexes only slightly. Emission and excitation fluorescence spectra of the solubilized membranes resemble spectra of trimers of PSII light-harvesting complexes (LHCII). Despite a disrupted excitation energy transfer from LHCII to PSII antenna core in solubilized membranes, energy transfer from Chl b and carotenoids to emission forms of Chl a within LHCII trimers remained effective.     

Moderate heat stress reduces the pH component of the transthylakoid proton motive force in light-adapted, intact tobacco leaves

We measured the Δ Ψ and ΔpH components of the transthylakoid proton motive force ( pmf ) in light-adapted, intact tobacco leaves in response to moderate heat. The Δ Ψ causes an electrochromic shift (ECS) in carotenoid absorbance spectra. The light–dark difference spectrum has a peak at 518 nm and the two components of the pmf were separated by following the ECS for 25 s after turning the light off. The ECS signal was deconvoluted by subtracting the effects of zeaxanthin formation (peak at 505 nm) and the qE-related absorbance changes (peak at 535 nm) from a signal measured at 520 nm. Heat reduced ΔpH while Δ Ψ slightly increased. Elevated temperature accelerated ECS decay kinetics likely reflecting heat-induced increases in proton conductance and ion movement. Energy-dependent quenching (qE) was reduced by heat. However, the reduction of qE was less than expected given the loss of ΔpH. Zeaxanthin did not increase with heat in light-adapted leaves but it was higher than would be predicted given the reduced ΔpH found at high temperature. The results indicate that moderate heat stress can have very large effects on thylakoid reactions.     

Formation of superoxide anion and carbon-centered radicals by photosystem II under high light and heat stress—EPR spin-trapping study

In this study, electron paramagnetic resonance spin-trapping spectroscopy was used to study the light-induced production of superoxide anion (O₂ ^?-) and carbon-centered (R^?) radicals by Photosystem II (PSII). It is evidenced here that exposure of PSII membranes to high light (2,000 μmol photons m^?2 s^?1) or heat (47 °C) treatments prior to the illumination suppressed O₂ ^?- production, while R^? was formed. Formation of R^? in the both high light- and heat-treated PSII membranes was enhanced by DCMU. Removal of molecular oxygen by glucose/glucose oxidase/catalase system and O₂ ^?- scavenging by exogenous superoxide dismutase completely suppressed carbon-centered radical formation. It is proposed here that the oxidation of polyunsaturated fatty acids and amino acids by O₂ ^?- on the electron acceptor side of PSII results in the formation of R^?, known to initiate a cascade reaction leading to the lipid peroxidation and protein degradation, respectively.     

Effect of partial or complete elimination of light‐harvesting complexes on the surface electric properties and the functions of cyanobacterial photosynthetic membranes

Influence of the modification of the cyanobacterial light‐harvesting complex [i.e. phycobilisomes (PBS)] on the surface electric properties and the functions of photosynthetic membranes was investigated. We used four PBS mutant strains of Synechocystis sp. PCC6803 as follows: PAL (PBS‐less), CK (phycocyanin‐less), BE (PSII‐PBS‐less) and PSI‐less/apcE^? (PSI‐less with detached PBS). Modifications of the PBS content lead to changes in the cell morphology and surface electric properties of the thylakoid membranes as well as in their functions, such as photosynthetic oxygen‐evolving activity, P700 kinetics and energy transfer between the pigment–protein complexes. Data reveal that the complete elimination of PBS in the PAL mutant causes a slight decrease in the electric dipole moments of the thylakoid membranes, whereas significant perturbations of the surface charges were registered in the membranes without assembled PBS–PSII macrocomplex (BE mutant) or PSI complex (PSI‐less mutant). These observations correlate with the detected alterations in the membrane structural organization. Using a polarographic oxygen rate electrode, we showed that the ratio of the fast to the slow oxygen‐evolving PSII centers depends on the partial or complete elimination of light‐harvesting complexes, as the slow operating PSII centers dominate in the PBS‐less mutant and in the mutant with detached PBS.     

Mechanistic differences in photoinhibition of sun and shade plants

Leaf discs of the shade plant Tradescantia albiflora Kunth grown at 50 μmol · m^?2 · s^?1, and the facultative sun/shade plant Pisum sativum L. grown at 50 or 300 μmol · m^?2, s^?1, were photoinhibited for 4 h in 1700 μmol photons m^?2 · s^?1 at 22° C. The effects of photoinhibition on the following parameters were studied: i) photosystem II (PSII) function; ii) amount of D1 protein in the PSII reaction centre; iii) dependence of photoinhibition and its recovery on chloroplast-encoded protein synthesis; and, iv) the sensitivity of photosynthesis to photoinhibition in the presence or absence of the carotenoid zeaxanthin. We show that: i) despite different sensitivities to photoinhibition, photoinhibition in all three plants occurred at the reaction centre of PSII; ii) there was no correlation between the extent of photoinhibition and the degradation of the D1 protein; iii) the susceptibility to photoinhibition by blockage of chloroplas-tencoded protein synthesis was much less in shade plants than in plants acclimated to higher light; and iv) inhibition of zeaxanthin formation increased the sensitivity to photoinhibition in pea, but not in the shade plant Tradescantia. We suggest that there are mechanistic differences in photoinhibition of sun and shade plants. In sun plants, an active repair cycle of PSII replaces photoinhibited reaction centres with photochemically active ones, thereby conferring partial protection against photoinhibition. However, in shade plants, this repair cycle is less important for protection against photoinhibition; instead, photoinhibited PSII reaction centres may confer, as they accumulate, increased protection of the remaining connected, functional PSII centres by controlled, nonphotochemical dissipation of excess excitation energy.     

Acclimation of photosystem II to high temperature in a suspension culture of soybean (Glycine max) cells requires proteins that are associated with the thylakoid membrane

In a study of the responses of photosystem II (PSII) to high temperature in suspension-cultured cells of soybean (Glycine max L. Merr.), we found that high temperatures inactivated PSII via two distinct pathways. Inactivation of PSII by moderately high temperatures, such as 41°C, was reversed upon transfer of cells to 25°C. The recovery of PSII required light, but not the synthesis of proteins de novo. By contrast, temperatures higher than 45°C inactivated PSII irreversibly. An increase in the growth temperature from 25 to 35°C resulted in an upward shift of 3°C in the profile of the heat-induced inactivation of PSII, which indicated that the thermal stability of PSII had been enhanced. This acclimative response was reflected by the properties of isolated thylakoid membranes: PSII in thylakoid membranes from cells that had been grown at 35°C exhibited greater thermal stability than that from cells grown at 25°C. Disruption of the vesicular structure of thylakoid membranes with 0.05% Triton X-100 decreased the thermal stability of PSII to a similar level in both types of thylakoid membrane. Proteins released by Triton X-100 from thylakoid membranes from cells grown at 35°C were able to increase the thermal stability of Triton-treated thylakoid membranes. These observations suggest that proteins that are associated with thylakoid membranes might be involved in the enhancement of the thermal stability of PSII.     

Slow degradation of the d1 protein is related to the susceptibility of low-light-grown pumpkin plants to photoinhibition

Photoinhibition of photosystem II (PSII) electron transport and subsequent degradation of the D1 protein were studied in pumpkin (Cucurbita pepo L.) leaves developed under high (1000 μmol m⁻² s⁻¹) and low (80 μmol m⁻² s⁻¹) photon flux densities. The low-light leaves were more susceptible to high light. This difference was greatly diminished when illumination was performed in the presence of chloramphenicol, indicating that a poor capacity to repair photodamaged PSII centers is decisive in the susceptibility of low-light leaves to photoinhibition. In fact, the first phases of the repair cycle, degradation and removal of photodamaged D1 protein from the reaction center complex, occurred slowly in low-light leaves, whereas in high-light leaves the degradation of the D1 protein more readily followed photoinhibition of PSII electron transport. A modified form of the D1 protein, with slightly slower electrophoretic mobility than the original D1, accumulated in the appressed thylakoid membranes of low-light leaves during illumination and was subsequently degraded only slowly.     

Effects of electric field on the photocycle of bacteriorhodopsin

The photoconversion of bacteriorhodopsin and the effects of an applied electric field (5 · 10⁷ V · m^?1) were studied in dry films of purple membranes from Halobacterium halobium. The electric field was found to cause at least two different effects: (1) it blocks in part the formation of the batho-bacteriorhodopsin (K), most probably due to electrically-induced dark transition of some bacteriorhodopsin molecules into the photochemically inactive form; (2) it decreases the rate of the intermediate M decay, the rise time of the M formation being unaffected by electric field. The observed phenomena may suggest a feedback control mechanism for the regulation of the bacteriorhodopsin photocycle in purple membranes.     

Photosynthesis and photoprotection in mangroves under field conditions

Net CO₂ exchange and in vivo chlorophyll fluorescence were studied in mangrove (Rhizophora stylosa) leaves at a field site in Western Australia, and leaf samples were collected for the analysis of enzymes and substrates potentially involved in anti-oxidant photoprotection. Photosynthesis saturated at 900 μmol quanta m^?2 s^?1 and at no more than 7.5 μmol CO₂ m^?2 s^?1. However, fluorescence analysis indicated no chronic photoinhibition: F_v:F_m was 0.8 shortly after sunset, and quantum efficiencies of PSII were high up to 500 μmol quanta m^?2 s^?1. Electron flow through PSII was more than 3 times higher than electron consumption through Calvin cycle activity, however, even with photorespiration and temperature-dependent Rubisco specificities taken into account. Acknowledging the growing body of literature attributing a role to antioxidant systems in photoprotection, we also assayed the activities of superoxide dismutase (SOD) and several enzymes potentially involved in H₂O₂ metabolism. Their levels of maximal potential activity were compared with those in greenhouse-grown mangroves (R. mangle), and growth chamber-grown peas. Monodehydroascorbate reductase activities were similar in all species, and glutathione reductase was lower, and ascorbate peroxidase ～40% higher, in the mangroves. SOD activities in field-grown mangroves were more than 40 times those in peas. Our results support the hypothesis that O₂ may be a significant sink for photochemically derived electrons under field conditions, and suggest an important role for O₂^? scavenging in photoprotection. However, when relative patterns are compared between species, imbalances between SOD and the other enzymes in the mangroves suggest that more components of the system (e.g. phenolics or peroxidases) are yet to be identified.     

Essential role of the PSI–LHCII supercomplex in photosystem acclimation to light and/or heat conditions by state transitions

Light and temperature affect state transitions through changes in the plastoquinone (PQ) redox state in photosynthetic organisms. We demonstrated that light and/or heat treatment induced preferential photosystem (PS) I excitation by binding light-harvesting complex II (LHCII) proteins. The photosystem of wheat was in state 1 after dark overnight treatment, wherein PQ was oxidized and most of LHCII was not bound to PSI. At the onset of the light treatment [25 °C in the light (100 µmol photons m^?2 s^?1)], two major LHCIIs, Lhcb1 and Lhcb2 were phosphorylated, and the PSI–LHCII supercomplex formed within 5 min, which coincided with an increase in the PQ oxidation rate. Heat treatment at 40 °C of light-adapted wheat led to further LHCII protein phosphorylation of, resultant cyclic electron flow promotion, which was accompanied by ultrafast excitation of PSI and structural changes of thylakoid membranes, thereby protecting PSII from heat damage. These results suggest that LHCIIs are required for the functionality of wheat plant PSI, as it keeps PQ oxidized by regulating photochemical electron flow, thereby helping acclimation to environmental changes.     

ACCLIMATION OF SYNECHOCOCCUS STRAIN WH7803 TO AMBIENT CO2 CONCENTRATION AND TO ELEVATED LIGHT INTENSITY1

A CO₂ concentrating mechanism has been identified in the phycoerythrin-possessing Synechococcus sp. WH7803 and has been observed to be severely inhibited by short exposure to elevated light intensities. A light treatment of 300–2000 μmol quanta·m^?2·s^?1 resulted in a considerable decay in the variable fluorescence of PSII with time, suggesting decreased efficiency of energy transfer from the phycobilisomes, direct damage to the reaction center II, or both. Measurements of the activity of PSII and changes in fluorescence emission spectra during a light treatment of 1000 μmol quanta·m^?2·s^?1 indicated considerable reduction in the energy flow from the phycocyanin to the phycobilisome terminal acceptor and chlorophyll a. Consequently, whereas the maximal photosynthetic rate, at saturating light and Co₂ concentration, was hardly affected by a light treatment of 1000 μmol quanta·m^?2·s^?1 for 2 h, the light intensity required to reach that maximum increased with the duration of the light treatment.     

Photon irradiance required to support optimal growth and interrelations between irradiance and pigment composition in the green alga Haematococcus pluvialis

The aim of the work was to find the optimal photon irradiance for the growth of green cells of Haematococcus pluvialis and to study the interrelations between changes in photochemical parameters and pigment composition in cells exposed to photon irradiances between 50 and 600?µmol?m^?2?s^?1 and a light:dark cycle of 12:12?h. Productivity of cultures increased with irradiance. However, the rate of increase was higher in the range 50–200?µmol?^?2?s^?1. The carotenoid content increased with increasing irradiance, while the chlorophyll content decreased. The maximum quantum yield of PSII (F_v/F_m) gradually declined from 0.76 at the lowest irradiance of 50?µmol?^?2?s^?1 to 0.66 at 600?µmol?^?2?s^?1. Photosynthetic activity showed a drop at the end of the light period, but recovered fully during the following dark phase. A steep increase in non-photochemical quenching was observed when cultures were grown at irradiances above 200?µmol?^?2?s^?1. A sharp increase in the content of secondary carotenoids also occurred above 200?µmol?m^?2?s^?1. According to our results, with H. pluvialis green cells grown in a 5-cm light path device, 200?µmol?^?2?s^?1 was optimal for growth, and represented a threshold above which important changes in both photochemical parameters and pigment composition occurred.     

Relationship between chloroplast structure and O2 evolution rate of leaf discs in plants from different biotopes in South Finland

Abstract The chloroplast ultrastructure, especially the thylakoid organization, the polypeptide composition of the thylakoid membranes and photosynthetic O₂ evolution rate, chlorophyll (Chl) content and Chi a/b ratio were studied in leaves of nine plants growing in contrasting biotopes in the wild in South Finland. All the measurements were made at the beginning of the period of main growth on leaves approaching full expansion, when the CO₂-saturated O₂ evolution rate (measured at 20°C and 1500 μmol photons m^?2s^?1) was at a maximum, ranging from 19.2 to 6.9 μmol O₂ cm^?2 h^?1. Among the species, the Chi a/b ratio varied between 3.75 and 2.71. In the mesophyll chloroplasts, the ratio of the total length of appressed to non-appressed thylakoid membranes varied between 1.07 and 1.79, the number of partitions per granum varied between 2.8 and 12.0 and the grana area between 21 and 42% of the chloroplast area. There was a significant relationship between the rate of O₂ evolution of the leaf discs and the thylakoid organization in the mesophyll chloroplasts. The higher the O₂ evolution rate, the lower was the ratio of the total length of appressed to non-appressed thylakoid membranes and also the lower the grana area. Although the relationship of the photosynthetic rate with the Chi content and the Chi a/b ratio of the leaves was not as clear, a significant negative correlation existed between the Chi a/b ratio and the ratio of appressed to non-appressed thylakoid membranes, indicating lateral heterogeneity in the distribution of different Chl- protein complexes.     

Photochemical activity and the structure of chloroplasts in <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis> L. mutants deficient in phytochrome A and B

The reduced content of photoreceptors, such as phytochromes, can decrease the efficiency of photosynthesis and activity of the photosystem II (PSII). For the confirmation of this hypothesis, the effect of deficiency in both phytochromes (Phy) A and B (double mutant, DM) in 7–27-day-old Arabidopsis thaliana plants on the photosynthetic activity was studied in absence and presence of UV-A radiation as a stress factor. The DM with reduced content of apoproteins of PhyA and PhyB and wild type (WT) plants with were grown in white and red light (WL and RL, respectively) of high (130 μmol quanta m^?2 s^?1) and low (40 μmol quanta m^?2 s^?1) intensity. For DM and WT grown in WL, no notable difference in the photochemical activity of PSII was observed. However, the resistance of the photosynthetic apparatus (PA) to UV-A and the rate of photosynthesis under light saturation were lower in the DM compared to those in the WT. Growth in RL, when the photoreceptors of blue light—cryptochromes—are inactive, resulted in the significant decrease of the photochemical activity of PSII in DM compared to that in WT including amounts of Q_B-non-reducing complexes of PSII and noticeable enhancement of thermal dissipation of absorbed light energy. In addition, marked distortion of the thylakoid membrane structure was observed for DM grown in RL. It is suggested that not only PhyA and PhyB but also cryptochromes are necessary for normal functioning of the PA and formation of the mechanisms of its resistance to UV-radiation.     

Alteration of thylakoid membrane structure in Brassica rapa ssp. oleifera during ageing in high and low light

Abstract Alterations in the composition and structure of thylakoids were studied in Brassica rapa ssp. oleifera grown under high and low irradiance (800 μmol m^?2 s^?1 and 80 μmol m^?2 s^?1). During ageing, both high and low light induced a decrease in total protein particle density and in the relative amount of 80–90 Å cytochrome b6/f and 90–100 Å ATP-synthetase. The density of PSII complexes in stacked (EFs) and unstacked (EFu) thylakoids also decreased. In high light, a shift was noted towards smaller PSII complexes in the EFs face with decreasing attached antenna complex CP29, but the relative amount of the antenna chlorophyll a-protein complexes of photosystem II (CPa) remained stable. In contrast, the proportion of peripheral LHCH on the PFs face and the density of PFs particles increased together with an increase in grana size. In low light, a shift occurred towards larger PSII complexes on the EFs face, along with a decrease in the proportion of CPa complexes and the PFs particle density (peripheral LHCH), though a marked increase was observed in the proportion of chlorophyll a/b-protein complexes in SDS-PAGE. The amount of photosystem I in green gel remained fairly stable, although the density of PFu particles (including PSI) increased in low and slightly diminished in high light. The results indicate that the organization of thylakoid components depends strongly on the light conditions and stage of development.     

Changes in Unsaturated Levels of Fatty Acids in Thylakoid PSII Membrane Lipids During Chilling-induced Resistance in Rice

Temperature is one of the abiotic factors limiting growth and productivity of plants. In the present work, the effect of low non‐freezing temperature, as an inducer of “chilling resistance”, was studied in three cultivars of rice (Oryza sativa L.), japonica cv. 9516 (j‐9516), the two parental lines of superhigh‐yield hybrid rice between subspecies, Peiai/E32 (ji‐PE), and the traditional indica hybrid rice Shanyou 63 (i‐SY63). Leaves of chill‐treated rice showed chilling‐induced resistance, as an increase of their low‐temperature tolerance was measured using chlorophyll fluorescence measurements, revealing a change in photosystem II (PSII) efficiency. After 5 d of exposure to 11°C under low light (100 μmol m^‐2 s^‐1), levels of unsaturated fatty acids in PSII thylakoid membrane lipids decreased during the initial 1‐2 d, then increased slowly and reached 99.2%, 95.3% and 90.1% of the initial value (0 d) in j‐9516, ji‐PE and i‐SY63, respectively, on the fifth day. However, under medium light (600 μmol m^‐2 s^‐1), all cultivars experienced similar substantial photoinhibition, which approached steady state levels after a decline in levels of unsaturated fatty acids in PSII thylakoid membrane lipids to about 57.1%, 53.8% and 44.5% of the initial values (0 d) in j‐9516, ji‐PE and I‐SY63 on the fifth day. Under either chilling‐induced resistance (the former) or low temperature photoinhibition (the latter) conditions, the changes of other physiological parameters such as D1 protein contents, electron transport activities of PSII (ETA), F_v/F_m, xanthophyl cycle activities expressed by DES (deepoxide state) were consistent with that of levels of unsaturated fatty acids in PSII thylakoid membrane lipids. So there were negative correlations between saturated levels of fatty acids (16:1(3t), 16:0, 18:0), especially the 16:1(3t) fatty acid on thylakoid membrane and other physiological parameters, such as D1 protein contents, ETA and (A+Z)/(A+V+Z). A specific role of desaturation of fatty acids and the photoprotective pigments of the xanthophyl cycle, leading to an acclimation response in thylakoid membrane lipids may be involved. We conclude that chilling‐induced resistance is accelerated by the unsaturation of thylakoid membranes, and the ability of rice plants to cold‐harden can be enhanced by genetic engineering.     

The effects of external electric field on the electron transport system of spinach chloroplasts

Since the thylakoid membranes of an active chloroplast are constantly exposed to the electric fields generated by the electron transport system inside the membranes, we have studied the effects of pretreating chloroplasts of spinach ( Spinacia oleracea L.) leaves with an external AC (alternating current) electric field on their electron transport system. It was found that a few minutes electric field pretreatment (333 V cm^-1 across chloroplast samples), especially at low frequency, irreversibly inhibited the activity of photosystem II (PSII), but under certain conditions, stimulated that of photosystem I (PSI). From the measurements of fluorescence from PSII, we ascribe the inhibition to a lesion close to its reaction center P680, leading to increased dissipation of excitation energy to heat. The effect on PSI was investigated by the reduction of its reaction center, P700 by various artificial donors. We suggest that the stimulative effect can be attributed to a positive shift of the surface charge density of thylakoid membranes that brings about an increase in the accessibility of exogenous electronegative donors.     

Effect of action potential on photosynthesis and spatially distributed H+ fluxes in cells and Chloroplasts of Chara corallina

When exposed to light, the cells of characean algae produce intermittent regions of H⁺ extrusion and H⁺ absorption, featuring different photosynthetic activities. Methods for local measurements of outer pH, O₂ content, and photochemical activity of photosystem II (PSII) were applied to examine microscopic regions of Chara coralline Klein ex Willd. internodes. The results show that the functional spatial heterogeneity of these excitable cells is controlled not only by light but also by electric excitation of the plasma membrane. Generation of a single action potential (AP) induced a reversible transition to the state with homogenous pH distribution and had different effects on photosynthesis in cell regions producing alkaline and acid zones. The effective quantum yield of PSII primary processes and the maximal chlorophyll fluorescence decreased after AP in the alkaline cell regions but were almost unaffected in the acidic cell regions. The suppression of photosynthesis after AP was also evident in the decrease of photosynthetic O₂ evolution. The results provide evidence that electric signals arising at the plasmalemma are transmitted to the level of thylakoid membranes. The effects of electric excitation on fluorescence and the quantum yield of PSII photochemistry were best pronounced at low light intensities and low level of nonphotochemical quenching. The sensitivity of chlorophyll fluorescence in resting and excited cells to light intensity and protonophores indicates that the AP-induced fluorescence changes derive from the increase in pH gradient at the thylakoid membrane. The temporal elimination of alkaline zones and inhibition of photosynthesis apparently arise from parallel operational sequences that have a common initial stage. A possible role of cytosolic Ca²⁺ rise in the mechanism of photosynthesis suppression after electric excitation of the plasma membrane is discussed.     

Electrochromic absorbance changes of photosynthetic pigments in Rhodopseudomonas sphaeroides II. Analysis of the band shifts of carotenoid and bacteriochlorophyll

An analysis was made of the changes of pigment absorption upon illumination of chromatophores of Rhodopseudomonas sphaeroides at ?35 °C, described in the preceding paper (de Grooth, B. G. and Amesz, J. (1977) Biochim. Biophys. Acta 462, 237–246). Comparison of the light-induced difference spectra in the carotenoid region obtained without additions, and in the presence of N-methylphenazonium methosulphate and ascorbate as donor-acceptor system showed that the latter spectrum was not only about 10 times larger in amplitude, but also red-shifted with respect to the first one. Together with the shape of the difference spectrum, this indicated that the spectrum obtained in the presence of a donor-acceptor system is due to an electrochromic shift of the absorption spectrum of a carotenoid by a few nm towards longer wavelength, caused by a delocalized potential across the chromatophore membrane. The results of an analysis of the kinetics of the absorbance changes near the zero points of the spectrum were in quantitative agreement with the extent of the red shift and indicated a shift of 0.25 nm for a single electron transfer per reaction center, and shifts of up to 4 nm when the electron transport is stimulated by a donor-acceptor system. For bacteriochlorophyll B-850 the shift is three times smaller.Analysis of the overall absorption spectrum showed that there are at least two pools of carotenoid. The carotenoid that shows electrochromism has absorption bands at 452, 481 and 515 nm, and comprises about one-third of the total carotenoid present; the remaining pool absorbs at about 7 nm shorter wavelength and does not show an electrochromic response to illumination. Both pools presumably consist of spheroidene; the differences in band location may be explained by the assumption that only the first pool is subjected to a local electric field which induces an electric dipole even at zero membrane potential. Similar results were obtained at room temperature and with a mutant of Rps. sphaeroides (G1C)-containing neurosporene.