Evidence against proton gradient formation being the cause of chlorophyll fluorescence quenching by N-methylphenazonium methosulfate.

In strong illumination, 3-(3, 4-dichlorophenyl)-1,1-dimethylurea (DCMU)-poisoned chloroplasts exhibit a high yield of chlorophyll fluorescence while P-700 turnover, proton uptake, and phosphorylation are inhibited and a pH gradient is undectectable. When 10muM N-methylphenazonium methosulfate (PMS) is included, the fluorescence yield in light is substantially reduced, and when 100 muM ascorbate is also included, the yield is diminished approximately to the level in darkness. Only very slight increases in P-700 turnover and proton uptake (but no detectable pH gradient) accompany the fluorescence yield decline. When 10muM PMS and 15 mM ascorbate are added to poisoned chloroplasts (the oxygen concentration being greatly reduced), P-700 turnover, proton uptake, the pH gradient and phosphorylation all reach high levels. In this case, the yield of chlorophyll fluorescence is low and is the same in both light and dark. Further addition of an uncoupler eliminates proton uptake, the pH gradient and phosphorylation but does not significantly elevate the fluorescence yield. From these observations we suggest that, in DCMU-poisoned chloroplasts, the fluorescence quenching with PMS occurrs by a mechanism unrelated to the generation of a phosphyorylation potential. With chloroplasts unpoisoned by DCMU, PMS quenches fluorescence and considerably stimulates proton uptake, the pH gradient and phosphorylation. However, in this case, PMS serves to restore net electron transport.     

The Metabolism of Oat Leaves during Senescence: III. The Senescence of Isolated Chloroplasts

The changes in chlorophyll and protein in senescing chloroplasts isolated from the first leaves of 7-day-old oat (Avena sativa) seedlings have been investigated. In darkness the chlorophyll in these plastids is highly stable, losing only 5 to 10% of its content after 7 days at 26 C. This result contrasts with the behavior of chlorophyll in intact leaves, in which about 80% of the pigment would have disappeared in that time. The protein is less stable than the chlorophyll, though more stable than in the leaf; probably a small amount of protease is present in the plastids. Some protein is also being synthesized in the chloroplasts along with its breakdown; gains of up to 38% in protein and 13% in chlorophyll were observed under different conditions. l-Serine, which actively promotes senescence in the leaf, has only a very slight effect on the chloroplasts, and kinetin antagonizes it. Kinetin also has a small but significant effect in preserving the protein from breakdown. Acid pH somewhat promotes the breakdown, both of chlorophyll and protein. A loss of chlorophyll and protein comparable to that occurring in the senescence of the leaf could not be induced in the chloroplasts by suspending them in malate, in cytoplasmic extract, or in any of a number of enzymes tested alone. Incubation with a mixture of four enzymes was the only treatment which approximated the senescent process in the leaf, causing 34% loss of chlorophyll at pH 5 and 40% loss of protein at pH 7.4, both in 72 hours.In white light, the chlorophyll and the carotenoids, but not the protein, disappear rapidly. This disappearance was shown to be prevented in an atmosphere of nitrogen or in air by a number of reducing agents, of which ascorbic acid was the most effective. It is, therefore, ascribed to photooxidation rather than to normal senescence.     

Luminescence Induced in Intact Leaves and Isolated Chloroplasts by Alcohols and Other Substances

Elodea leaves and spinach chloroplasts emit red light when treated with alcohols or certain other solvents for chlorophyll. The intensity of the light, the lag phase and the threshold concentration vary considerably between different alcohols. Light emission from a leaf starts a few seconds to a few minutes after the addition of alcohol to the medium, reaches a maximum after 2–45 minutes (sometimes more) and then continues for many hours. Despite the faintness of the glow, the total number of photons given off from a leaf after addition of alcohol may exceed the number of photons given off from the same sample as long-lived afterglow after saturating irradiation with far-red light. The maximum yield of photons per chlorophyll molecule is a little more than 10^–5. The alcohol-induced luminescence is not influenced by a decrease in the oxygen tension to one fifth of the normal. Electron micrographs of treated leaves reveal that the thylakoid lipids contract to drops at the edges of the grana. Treatment of isolated chloroplasts with ethanol results in light emission when the concentration is high enough to dissolve the chlorophyll. It is estimated that the surface free energy in the thylakoid lipid-aqueous interface, due to ordinary interfacial tension, is large enough to account for the light emission observed when the interfaces contract or disappear.     

Chlorophyll fluorescence as a probe for the determination of the photo-induced proton gradient in isolated chloroplasts

Gramicidin D-treated chloroplasts show an acid-induced quenching of the chlorophyll fluorescence, which is composed of a reversible and irreversible part. The reversible quenching is analogous to the photo-induced quenching in coupled chloroplasts and can be taken to determine the light induced delta pH.     

Light-dependent changes of the Mg2+ concentration in the stroma in relation to the Mg2+ dependency of CO2 fixation in intact chloroplasts.

(1) Light-dependent changes of the Mg2+ content of thylakoid membranes were measured at pH 8.0 and compared with earlier measurements at pH 6.6. In a NaCl and KCl medium, the light-dependent decrease in the Mg2+ content of the thylakoid membranes at pH 8.0 is found to be 23 nmol Mg2+ per mg chlorophyll, whereas in a sorbitol medium it is 83 nmol Mg2+ per mg chlorophyll. (2) A light dependent increase in the Mg2+ content of the stroma was detected wjem chloroplasts were subjected to osmotic shock, amounting to 26 nmol/mg chlorophyll. Furthermore, a rapid and reversible light-dependent efflux of Mg2+ has been observed in intact chloroplasts when the divalent cation ionophore A 23 187 was added, indicating a light-dependent transfer of about 60 nmol of Mg2+ per mg chlorophyll from the thylakoid membranes to the stroma. (3) CO2 fixation, but not phosphoglycerate reduction, could be completely inhibited when A 23 187 was added to intact chloroplasts in the absence of external Mg2+. If Mg2+ was then added to the medium, CO2 fixation was restored. Half of the maximal restoration was achieved with about 0.2 mM Mg2+, which is calculated to reflect a Mg2+ concentration in the stroma of 1.2 mM. The further addition of Ca2+ strongly inhibits CO2 fixation. (4) The results suggest that illumination of intact chloroplasts causes an increase in the Mg2+ concentration of 1-3 mM in the stroma. Compared to the total Mg2+ content of chloroplasts, this increase is very low, but it appears to be high enough to have a possible function in the light regulation of CO2 fixation.     

Effects of High K+ and Alkaline pH on Ultrastructure of Dunaliella salina Chloroplasts

It was reported that the growth of Dunaliella salina Teod. cultured in medium containing 1 mol/L NaC1 was almost completely inhibited by the addition of 100 mmol/L KC1. The high K+ (100 mmol/L KC1) treatment also significantly inhibited the photosynthetic rate of D. salina and decreased chlorophyll contents in algae. This study focuses on possible effects of high K+ or alkaline pH on the ultrastructural change of chloroplasts in D. salina. After D. salina was cultured in a medium containing 100 n,anol/L KC1 or in a medium with alkaline pH for 8 to 10 days, dramatic ultrastructural changes occurred in the chloroplasts including thylakoid swelling, volume increase of chloroplast, and significant accumulation of starch grains in chloroplasts. The results are consistent with our previous report indicating that the ultrastmctuml changes in chloroplast under high K + or alkaline pH may lead to an inhibitory effects on photosynthesis and overall growth of D. salina.     

Evidence against proton gradient formation being the cause of chlorophyll fluorescence quenching by N-methylphenazonium methosulfate

In strong illumination, 3-(3,4-dichlorophenyl)-1, 1-dimethylurea (DCMU)-poisoned chloroplasts exhibit a high yield of chlorophyll fluorescence while $\text{P-700}$ turnover, proton uptake, and phosphorylation are inhibited and a pH gradient is undetectable. When 10 μM $\text{N-}\text{methylphenazonium}$ methosulfate (PMS) is included, the fluorescence yield in light is substantially reduced, and when 100 μM ascorbate is also included, the yield is diminished approximately to the level in darkness. Only very slight increases in $\text{P-700}$ turnover and proton uptake (but no detectable pH gradient) accompany the fluorescence yield decline.When 10 μM PMS and 15 mM ascorbate are added to poisoned chloroplasts (the oxygen concentration being greatly reduced), $\text{P-700}$ turnover, proton uptake, the pH gradient and phosphorylation all reach high levels. In this case, the yield of chlorophyll fluorescence is low and is the same in both light and dark. Further addition of an uncoupler eliminates proton uptake, the pH gradient and phosphorylation but does not significantly elevate the fluorescence yield. From these observations we suggest that, in DCMU-poisoned chloroplasts, the fluorescence quenching with PMS occurs by a mechanism unrelated to the generation of a phosphorylation potential.With chloroplasts unpoisoned by DCMU, PMS quenches fluorescence and considerably stimulates proton uptake, the pH gradient and phosphorylation. However, in this case, PMS serves to restore net electron transport.     

High Light-Induced Unstacking of Thylakoids and Randomization of Pigment-Protein Complexes of Chloroplast

Using absorbance changes at 540 nm and the per cent chlorophyll in 10K pellet after digitonin fractionation as the criteria of thylakoid stacking, high light induced unstacking of thylakoids and randomization of pigment protein complexes of chloroplast was characterized. Cation-induced increase in absorbance at 540 nm was abolished when chloroplasts were exposed to high light. Also, the high light induced decrease in absorbance at 540 nm was more pronounced In low salt chloroplasts as compared to the decrease in 540 nm absorbance in chloroplasts isolated in high salt. The decrease In the fraction of chlorophyll in digitonin fractionated 10K pellet was much more than the decrease in chlorophyll in 10K pellet from high salt chloroplasts exposed to high light These results suggest that high light causes unstacking of thylakoids.     

Light-dependent changes of the Mg concentration in the stroma in relation to the Mg dependency of CO2 fixation in intact chloroplasts

(1) Light-dependent changes of the Mg²⁺ content of thylakoid membranes were measured at pH 8.0 and compared with earlier measurements at pH 6.6. In a NaCl and KCl medium, the light-dependent decrease in the Mg²⁺ content of the thylakoid membranes at pH 8.0 is found to be 23 nmol Mg²⁺ per mg chlorophyll, whereas in a sorbitol medium it is 83 nmol Mg₂₊ per mg chlorophyll.

(2) A light dependent increase in the Mg²⁺ content of the stroma was detected when chloroplasts were subjected to osmotic shock, amounting to 26 nmol/mg chlorophyll. Furthermore, a rapid and reversible light-dependent efflux of Mg²⁺ has been observed in intact chloroplasts when the divalent cation ionophore A 23 187 was added, indicating a light-dependent transfer of about 60 nmol of Mg²⁺ per mg chlorophyll from the thylakoid membranes to the stroma.

(3) CO₂ fixation, but not phosphoglycerate reduction, could be completely inhibited when A 23 187 was added to intact chloroplasts in the absence of external Mg²⁺. If Mg²⁺ was then added to the medium, CO₂ fixation was restored. Half of the maximal restoration was achieved with about 0.2 mM Mg²⁺, which is calculated to reflect a Mg²⁺ concentration in the stroma of 1.2 mM. The further addition of Ca²⁺ strongly inhibits CO₂ fixation.

(4) The results suggest that illumination of intact chloroplasts causes an increase in the Mg²⁺ concentration of 1–3 mM in the stroma. Compared to the total Mg²⁺ content of chloroplasts, this increase is very low, but it appears to be high enough to have a possible function in the light regulation of CO₂ fixation.     

The reduction kinetics of chlorophyll aI as an indicator for proton uptake between the light reactions in chloroplasts.

The flash-induced oxidation kinetics of the primary acceptor of light Reaction II (X-320) and the reduction kinetics of chlorophyll aI (P-700) after far-red preillumination have been studied with high time resolution in spinach chloroplasts. 1. The kinetics of chlorophyll aI exhibits a pronounced lag phase of 2--3 ms at the onset of reduction as would be expected for the final product of consecutive reactions. Because the oxidation of the plastoquinone pool is the rate-limiting step for the electron transport between the two light reactions, the lag indicates the maximal electron transfer time over all preceding reactions after light Reaction II. 2. The observation that the lag phase decreases with decreasing pH is evidence of an electron transfer step coupled to a proton uptake reaction. 3. Protonation of X-320 after reduction in the flash is excluded because a slight increase of the decay time is found at decreasing pH values. 4. The time course of plastohydroquinone formation is deduced from the first derivative of the reduction kinetics of chlorophyll aI. This approach covers those plastohydroquinone molecules being available to the electron carriers of System I via the rate-limiting step. Direct measurements of absorbance changes would not allow to discriminate between these and functionally different plastohydroquinone molecules. 5. The derived time course of plastohydroquinone at different pH gives evidence for an additional electron transfer step with a half time of about 1 ms following the proton uptake and preceding the rate-limiting step. It is tentatively attributed to the diffusion of neutral plastohydroquinone across the hydrophobic core of the thylkaloid membrane. 6. The lower limit of the rate constant for proton uptake by an electron carrier, consistent with the lag of chlorophyll aI reduction, is estimated as greater than 10(11) M-1s-1. The value is higher than that of the fastest diffusion controlled protonations of organic molecules in solution. Possible mechanisms of linear electron transport between light Reaction II and the rate-limiting oxidation of neutral plastohydroquinone are thoroughly discussed.     

Surface charges on chloroplast membranes as studied by particle electrophoresis.

1. Particle microelectrophoresis mobility studies have been conducted with chloroplast thylakoid membranes and with isolated intact chloroplasts. 2. The pH dependence of the electrophoretic mobility indicated that at pH values above 4.3 both membrane systems carry a net negative charge. 3. Chemical treatment of thylakoids has shown that neither the sugar residues of the galactolipids in the membrane nor the basic groups of the membrane proteins having pK values between 6 and 10 are exposed at the surface. 4. However, treatment with 1-ethyl-3(3-dimethylaminopropyl)carbodiimide, together with glycine methyl ester, neutralized the negative charges on the thylakoid membrane surface indicating the involvement of carboxyl groups which, because of their pH sensitivity, are likely to be the carboxyl groups of aspartic and glutamic acid residues. 5. The nature of the protein giving rise to the negative surface charges on the thylakoids is not known but is shown not to involve the coupling factor or the light harvesting chlorophyll a/chlorophyll b pigment . protein complex. 6. No significant effect of light was observed on the electrophoretic mobility of either thylakoids or intact chloroplasts. 7. The striking difference in the ability of divalent and monovalent cations to screen the surface charges was demonstrated and explained in terms of the Gouy-Chapman theory. 8. Calculations of the zeta-potentials for thylakoid membranes gave values for the charge density at the plane of shear to be in the region of one electronic charge per 1500--2000 A2. 9. The significance of the results is discussed in terms of cation distribution in chloroplasts and the effect of cations on photosynthetic phenomena.     

The Light Harvesting System of the Diatom Cyclotella cryptica. Isolation and Characterization of the Main Light Harvesting Complex and Evidence for the Existence of Minor Pigment Proteins*

The main chlorophyll a/c light harvesting complex of the diatom Cyclotella cryptica was isolated by sucrose density gradient centrifugation. It consisted of two polypeptides of Mrs 18000 and 22000. Both polypeptides and fragments thereof, obtained by formic acid treatment, were blocked at their N-ter-mini. An antiserum raised against the two subunits selectively immunolabeled the thylakoid within the chloroplasts. The subunits were nuclear encoded and could be immunoprecipitated from poly (A)⁺ RNA as precursor proteins in the Mr range of 20000 to 24000. The existence of minor chlorophyll protein complexes and their possible function in light climate adaptation processes was investigated in cells adapted to low light and high light conditions. Low light grown cells contained more fucoxanthin and less β-carotene relative to chlorophyll a than high light adapted cells. The xanthophyll cycle pigments diatoxanthin and diadinoxanthin increased five-fold relative to chlorophyll a under high light conditions. Western-immunoblotting experiments with antisera raised against several chlorophyll a/b and chlorophyll a/c antenna complexes demonstrated that, beside the dominating chlorophyll a/c light harvesting complex, minor antenna complexes might exist, which, in part, seem to react to the light climate applied.     

Removal of Mn from spinach chloroplasts by sodium cyanide and the binding of Mn2+ to Mn-depleted chloroplasts.

Manganese and copper were released from spinach chloroplasts by NaCN-treatment, though iron was not affected. The Hill reaction activity was also inhibited by this treatment, but was partially recovered by the addition of either Mn2+ or Cu2+, but not of Fe3+. The interaction of Mn2+ with manganese-depleted chloroplasts by NaCN-treatment was studied using 54Mn2+. A Scatchard plot shows the high and low affinity binding sites of Mn2+ on NaCN-treated chloroplast membrane; high affinity binding being specific for NaCN-treated chloroplast with a binding constant, KH, of 1.9 X 10(5) M-1, and a maximum binding number, NH, of 0.0016 g-atom per mole of chlorophyll. The low binding site was also found on untreated chloroplasts; its binding constant, KL, being 1.2 X 10(4) M-1, and its maximum binding number, NL, of 0.0112 g-atom per mole oc chlorophyll at pH 8.2 NH was proportional to the degree of the removal of Mn by NaCN-treatment and was constant at pH 4--9. NL markedly increased at a high pH with a midpoint of pH 7.9 indicating the exposure of a new, similar binding site. Light illumination partially inhibited the binding of Mn2+. Within 1 min in the dark the binding reaction reached equilibrium in the absence of pyrophosphate, however, 20 min were required to transform into pyrophosphate-resistant form. The pH dependence of the binding of Mn2+ with pKa 7.2 and the ineffectiveness of p-chloromercuribenzoate suggest the possible ligand of Mn2+ is the imidazole nitrogen of the histidine residue.     

Dissipation of the Proton Electrochemical Potential in Intact Chloroplasts (II. The pH Gradient Monitored by Cytochrome f Reduction Kinetics)

The potency of various uncouplers for collapsing the light-induced pH gradient across thylakoid membranes in intact chloroplasts was investigated by time-resolved optical spectroscopy. The thylakoid transmembrane pH gradient ([delta]pH) was monitored indirectly by measuring the rate of cytochrome (Cyt) f reduction following a light flash of sufficient duration to create a sizable [delta]pH. The results show that the rate of Cyt f reduction is controlled in part by the internal pH of the thylakoid inner aqueous space. At pH values from 6.5 to 8.0, the Cyt f reduction rate was maximal, whereas at lower pH values from 6.5 to 5.5 the reduction rate decreased to 25% of the maximal rate. The ability of three uncouplers, nigericin, carbonylcyanide m-chlorophenylhydrazone, and gramicidin, to accelerate the rate of Cyt f reduction was determined for intact chloroplasts isolated from spinach (Spinacia oleracea). The efficacy of the uncouplers for collapsing the [delta]pH was determined using the empirical relationship between the [delta]pH and the Cyt f reduction rate. For intact chloroplasts, nigericin was the most effective uncoupler, followed by carbonylcyanide m-chlorophenylhydrazone, which interacted strongly with bovine serum albumin. Gramicidin D, even at high gramicidin:chlorophyll ratios, did not completely collapse the pH gradient, probably because it partitions in the envelope membranes and does not enter the intact chloroplast.     

Acclimation of mesophyll and bundle sheath chloroplasts of maize to different irradiances during growth

The regulation by light of the photosynthetic apparatus, and composition of light-harvesting complexes in mesophyll and bundle sheath chloroplasts was investigated in maize. Leaf chlorophyll content, level of plastoquinone, PSI and PSII activities and Lhc polypeptide compositions were determined in plants grown under high, moderate and low irradiances. Photochemical efficiency of PSII, photochemical fluorescence quenching and non-photochemical fluorescence quenching over a range of actinic irradiances were also determined, using chlorophyll a fluorescence analysis. Acclimation of plants to different light conditions caused marked changes in light-harvesting complexes, LHCI and LHCII, and antenna complexes were also reorganized in these types of chloroplasts. The level of LHCII increased in plants grown in low light, even in agranal bundle sheath chloroplasts where the amount of PSII was strongly reduced. Irradiance also affected LHCI complex and the number of structural polypeptides, in this complex, generally decreased in chloroplasts from plants grown under lower light. Surprisingly moderate and low irradiances during growth do not affect the light reaction and fluorescence parameters of plants but generated differences in composition of light-harvesting complexes in chloroplasts. On the other hand, the changes in photosynthetic apparatus in plants acclimated to high light, resulted in a higher efficiency of photosynthesis. Based on these observations we propose that light acclimation to high light in maize is tightly coordinated adjustment of light reaction components/activity in both mesophyll and bundle sheath chloroplasts. Acclimation is concerned with balancing light utilization and level of the content of LHC complexes differently in both types of chloroplasts.     

pH dependent changes in the reactivity of the primary electron acceptor of System II in spinach chloroplasts to external oxidant and reductant

Changes in the rates of dark oxidation and reduction of the primary electron acceptor of System II by added oxidant and reductant were investigated by measuring the induction of chlorophyll fluorescence under moderate actinic light in 3-(3′,4′-dichlorophenyl)-1,1-dimethylurea-inhibited chloroplasts at pH values between 3.6 and 9.5. It was found that:

1. (1) The rate of dark oxidation of photoreduced primary acceptor was very slow at all the pH values tested without added electron acceptor.

2. (2) The rate was accelerated by the addition of ferricyanide in the whole pH range. It was dependent approximately on the 0.8th power of the ferricyanide concentration.

3. (3) The rate constant for the oxidation of the primary acceptor by ferricyanide was pH-dependent and became high at low pH. The value at pH 3.6 was more than 100 times that at pH 7.8.

4. (4) The pH-dependent change in the rate constant was almost reversible when the chloroplasts were suspended at the original pH after a large pH change (acid treatment).

5. (5) An addition of carbonylcyanide m-chlorophenylhydrazone or heavy metal chelators had little effect on the rate of dark oxidation of the primary acceptor by ferricyanide.

6. (6) The dark reduction of the primary acceptor by sodium dithionite also became faster at low pH.

From these results it is concluded that at low pH the primary acceptor of System II becomes accessible to the added hydrophilic reagents even in the presence of 3-(3′,4′-dichlorophenyl)-1,1-dimethylurea.     

Effects of ethanol on the interaction of photosynthetic processes in spinach chloroplasts

The interaction and coupling between photosynthetic processeswere studied in ethanol-treated chloroplasts (in the absenceof ethanol) or in the presence of ethanol. Light-induced H⁺uptake and photophosphorylation were suppressed and electrontransfer was enhanced by ethanol treatment or in the presenceof ethanol (10–25%). Dark recoveries of the H⁺ uptakeand the 515-nm absorbance change were accelerated by ethanol.The half-effective concentrations of ethanol for these processeswere higher in the ethanol-treated chloroplasts than when ethanolwas present in the reaction mixture. The maximum rate of electrontransfer in the ethanol-treated chloroplasts, which was at thesame level as that of the control with an uncoupler, was notaffected by uncouplers. The marked acceleration of recoveryof the 515-nm absorbance change by SCN- or valinomycin plusK⁺ in the untreated chloroplasts was much smaller in the ethanol-treatedchloroplasts or in the presence of ethanol. The ethanol-treatedpreparation had the same characteristics as those of the control,in chlorophyll fluorescence, light-intensity dependence of electrontransfer (compared with the control with an uncoupler), andsensitivity to sucrose osmolarity except for a slight increaseof the packed volume. Under the present conditions, the ethanoltreatment mainly induced an increase of permeability of thylakoidmembranes to ions. In chloroplasts treated with ethanol at various concentrations,the relationships of the uncoupler-stimulated part of electrontransfer with the dark half-recovery time of H⁺ uptake and withphotophosphorylation were linear. Logarithms of the photophosphorylationand the extent of H⁺ uptake had a linear relationship with aslope of about 3. This slope may indicate the stoichiometryof H⁺/ATP. (Received June 17, 1978; )     

Acclimation of mesophyll and bundle sheath chloroplasts of maize to different irradiances during growth

The regulation by light of the photosynthetic apparatus, and composition of light-harvesting complexes in mesophyll and bundle sheath chloroplasts was investigated in maize. Leaf chlorophyll content, level of plastoquinone, PSI and PSII activities and Lhc polypeptide compositions were determined in plants grown under high, moderate and low irradiances. Photochemical efficiency of PSII, photochemical fluorescence quenching and non-photochemical fluorescence quenching over a range of actinic irradiances were also determined, using chlorophyll a fluorescence analysis. Acclimation of plants to different light conditions caused marked changes in light-harvesting complexes, LHCI and LHCII, and antenna complexes were also reorganized in these types of chloroplasts. The level of LHCII increased in plants grown in low light, even in agranal bundle sheath chloroplasts where the amount of PSII was strongly reduced. Irradiance also affected LHCI complex and the number of structural polypeptides, in this complex, generally decreased in chloroplasts from plants grown under lower light. Surprisingly moderate and low irradiances during growth do not affect the light reaction and fluorescence parameters of plants but generated differences in composition of light-harvesting complexes in chloroplasts. On the other hand, the changes in photosynthetic apparatus in plants acclimated to high light, resulted in a higher efficiency of photosynthesis. Based on these observations we propose that light acclimation to high light in maize is tightly coordinated adjustment of light reaction components/activity in both mesophyll and bundle sheath chloroplasts. Acclimation is concerned with balancing light utilization and level of the content of LHC complexes differently in both types of chloroplasts.     

The differential scattering of circularly polarized light by chloroplasts and evaluation of their true circular dichroism

Chloroplasts isolated from pea leaves display an intense circular dichroism in the range 600 to 720nm. Circularly polarized light is also differentially scattered by chloroplasts, and this effect can be confused with circular dichroism. By using an instrumental modification it was possible to distinguish, and record separately, the ellipticities of the transmitted light (circular dichroism) and of the scattered light in the same c.d. instrument. By means of a light-scattering apparatus, the intensity of unpolarized light scattered by chloroplasts was measured as a function of wavelength and of angle. This measurement allowed the aforementioned ellipticities to be corrected for mutual interference. At a concentration of 4mug of chlorophyll/ml (the optimum practical concentration of chloroplasts at which there was no significant interaction of scattering and absorption effects) spectra of true circular dichroism (circular differential absorption) and circular differential scattering were obtained. The former showed maxima, positive at 688nm and negative at 676nm, with an intensity Deltatheta=8.3m degrees .litre.(mg of chlorophyll)(-1).cm(-1). The latter had a maximum at 683nm with an intensity of +47m degrees with respect to the solvent baseline; this value is independent of the concentration of chloroplasts in dilute suspensions. It is suggested that the intense circular dichroism of chloroplasts reflects specific chlorophyll-chlorophyll interactions in the light-harvesting pigment. The advantages of this method for determining the c.d. of scattering suspensions over those of other investigators are discussed.