Higher Electron Transport Rate Research Articles

Photosynthetic Electron Transport During Senescence of the Primary Leaves ofPhaseolus vulgarisL.: I. NON-CYCLIC ELECTRON TRANSPORT

Coupled, non-cyclic electron transport was measured for chloroplasts isolated from the primary leaves of Phaseolus vulgaris. Preparations from young, fully expanded leaves gave good rates of electron transport, but the rates obtained decreased by approximately 80% during leaf senescence. Higher rates of electron transport were recorded for chloroplasts isolated from primary leaves which had regreened following removal of the remainder of the shoot. With preparations from leaves of all ages, photophosphorylation was coupled to electron transport with a mean P/2e ratio of approximately 1.3. No evidence was obtained for inactivation of chloroplasts from older leaves during isolation or assay, and it is suggested that the decrease in rate of electron transport over the period of senescence, and its increase during regreening, were consequences of changes in the composition and physical properties of the thylakoid membrane which occur in vivo. The decrease in rate of non-cyclic electron transport may be important in limiting the rate of photosynthesis in the senescing leaves.

Photosynthetic electron transport in a cell-free preparation from the thermophilic blue-green alga Phormidium laminosum.

1. A cell-free preparation of membrane fragments was prepared from the thermophilic blue-green alga Phormidium laminosum by lysozyme treatment of the cells followed by osmotic shock to lyse the spheroplasts. The membrane fragments showed high rates of photosynthetic electron transport and O2 evolution (180-250 mumol of O2/h per mg of chlorophyll a with 2,6-dimethyl-1,4-benzoquinone as electron acceptor). O2-evolution activity was stable provided that cations (e.g. 10mM-Mg2+ or 100mM-Na+) or glycerol (25%, v/v) were present in the suspending medium. 2. The components of the electron-transport chain in P. laminosum were similar to those of other blue-green algae: the cells contained Pigment P700, plastocyanin, soluble high-potential cytochrome c-553, soluble low-potential cytochrome c-54 and membrane-bound cytochromes f, b-563 and b-559 (both low- and high-potential forms). The amounts and midpoint potentials of the membrane-bound cytochromes were similar to those in higher-plant chloroplasts. 3. Although O2 evolution in P. laminosum spheroplasts was resistant to high temperatures, thermal stability was not retained in the cell-free preparation. However, in contrast with higher plants, O2 evolution in P. laminosum membrane fragments was remarkably resistant to the non-ionic detergent Triton X-100.

Photoreactions of cytochromes in algal chloroplasts.

Light-induced cytochrome redox reactions were investigated with algal chloroplasts capable of high rates of electron transport coupled to phosphorylation. The electron-donor pool preceding photosystem I consists of membrane-bound cytochrome f-554.5 and a soluble cytochrome c-553; the latter replaces the function of plastocyanin of higher-plant chloroplasts. Both cytochromes are reduced by photosystem II and oxidized by photosystem I. A site of energy conservation precedes these c-type cytochromes. The data obtained with respect to the function of b-type cytochromes are comparable to those obtained with higher-plant chloroplasts. Cyclic electron transport is mediated by cytochrome b-563 in a photosystem-I-dependent reaction. In addition, cytochrome b-563 may be reduced by photosystem II, in accordance with recent findings with intact spinach chloroplasts. It therefore appears that cytochrome b-563 is a member of both cyclic and non-cyclic electron transport. In contrast to higher-plant chloroplasts, redox reactions of cytochrome b-559 are observable without any pretreatments. Cytochrome b-559, high-potential, is reduced by photosystem II through plastoquinone. In the presence of carbonyl cyanide m-chlorophenylhydrazone a cytochrome-b-559 oxidation by photosystem II is measured.

Consistently high rates of electron transport in chloroplasts isolated from hydroponically-grown spinach plants

Intact spinach chloroplasts were rapidly isolated from hydroponically grown spinach plants, immediately lysed and a number of different electron transport reactions assayed for the preparations. The observed rates of electron transport were considerably greater than those previously reported. It is suggested that the conditions under which plants are grown and the method of chloroplast isolation are important factors in determining the rate of electron transport in chloroplast preparations.

Quantum Requirements of Photosynthetic Electron Transport In Sitka Spruce from Different Light Environments

AbstractQuantum requirements of photosynthetic electron transport have been measured in shoots of Picea sitchensis (Bong.) Carr. (Sitka spruce) from different levels in a forest canopy and in shoots from plants grown in contrasting light environments in controlled environment chambers. Neutral density filters were used to obtain very low photon flux densities. The light absorbed by the chloroplast suspensions was calculated from measurements of the transmittance of the suspensions. The shoots from the top of the forest canopy (“sun” shoots) had lower quantum requirements for photosystems I and II than the shoots from the bottom of the forest canopy (“shade” shoots). High light grown plants and “sun” shoots had higher rates of electron transport at light saturation than low light grown plants and “shade” shoots. Thus a higher potential for electron transport was found to exist in “sun” shoots than in “shade” shoots at both high and low photon flux densities.

The role of plastidic cytochrome c in algal electron transport and photophosphorylation

By an improved isolation procedure chloroplasts could be obtained from the alga Bumilleriopsis filiformis (Xanthophyceae) which exhibited high electron transport rates tightly coupled to ATP formation. Uncouplers both stimulate electron transport and inhibit photophosphorylation. These chloroplasts retain almost all soluble cytochrome c-553 besides a membrane-bound cytochrome c-554.5 (= f-554.5). Sonification or iron deficiency removed the soluble cytochrome only with a concurrent decrease of electron transport from water to methyl viologen or to NADP and decreased non-cyclic and cyclic photophosphorylation. However, photosynthetic control and the P 2e ratios remain unaltered. In Bumilleriopsis, which apparently has no plastocyanin, the soluble cytochrome c-553 seemingly links electron transport between the bound cytochrome c and P-700.

Electron paramagnetic resonance studies of photosynthetic electron transport: Photoreduction of ferredoxin and membrane-bound iron-sulfur centers

Electron paramagnetic resonance spectrometry was used to investigate, at physiological temperatures, light-induced electron transport from membrane-bound iron-sulfur components (bound ferredoxin) to soluble ferredoxin and NADP(+) in membrane fragments (from the blue-green alga, Nostoc muscorum) that had high rates of electron transport from water to NADP(+) and from an artificial electron donor, reduced dichlorophenolindophenol (DCIPH(2)) to NADP(+). Illumination at 20 degrees resulted in the photoreduction of membrane-bound iron-sulfur centers A and B. Photoreduction by water gave electron paramagnetic resonance signals of both centers A and B; photoreduction by DCIPH(2) was found to generate a strong electron paramagnetic signal of only center B. When water was the reductant, the addition and photoreduction of soluble ferredoxin generated additional signals characteristics of soluble ferredoxin without causing a decrease in the amplitude of the signals due to centers A and B. The further addition of NADP(+) (and its photoreduction) greatly diminished signals due to the bound iron-sulfur centers and to soluble ferredoxin. An outflow of electrons from center B to soluble ferredoxin and NADP(+) was particularly pronounced when DCIPH(2) was the reductant. These observations provide the first evidence for a light-induced electron transport between membrane-bound iron-sulfur centers and ferredoxin-NADP(+). The relationship of these observations to current concepts of photosynthetic electron transport is discussed.

Effect of the Plastoquinone Antagonist Dibromothymoquinone on Electron Transport Reactions in Chloroplast Preparations

The effects of dibromothymoquinone (DBMIB) on electron transport in types A, B, C and D spinach and pea chloroplast preparations have been studied. DBMIB (1 x 10-6M) strongly inhibited electron transport to methyl viologen in all the types of chloroplasts, while electron transport to ferricyanide was inhibited by 60%. Envelope-free (type C) chloroplast preparations with poor photosynthetic control were inhibited more strongly than intact (types A and B) chloroplast preparations at low (< 5 x 10-7M) concentrations of DBMIB. Type C preparations with good photosynthetic control were less strongly inhibited except in the presence of uncoupler or in the absence of ADP. Above 1 x 10-6M DBMIB, inhibition of electron transport to ferricyanide became progressively less and, with type A chloroplast preparations, a large stimulation compared to the control level occurred. Phenylenediamine stimulated high rates of electron transport to ferricyanide in the presence of low concentrations of DBMIB. At higher DBMIB concentrations, the stimulation was completely reversed and envelope-free chloroplasts again showed greater sensitivity to DBMIB compared to intact chloroplasts. DBMIB appears to have a number of sites of interaction with the thylakoid membrane.

Effects of magnesium and chloride ions on light-induced electron transport in membrane fragments from a blue-green alga

The effects of magnesium and chloride ions on photosynthetic electron transport were investigated in membrane fragments of a blue-green alga, Nostoc muscorum (Strain 7119), noted for their stability and high rates of electron transport from water or reduced dichlorophenolindophenol to NADP +. Magnesium ions were required not only for light-induced electron transport from water to NADP + but also for protection in the dark of the integrity of the water-photooxidizing system (Photosystem II). Membrane fragments suspended in the dark in a medium lacking Mg 2+ lost the capacity to photoreduce NADP + with water on subsequent illumination. Chloride ions could substitute, but less effectively, for each of these two effects of magnesium ions. By contrast, the photoreduction of NADP + by DCIPH 2 was independent of Mg 2+ (or Cl −) for the protection of the electron transport system in the dark or during the light reaction proper. Furthermore, high concentrations of MgCl 2 produced a strong inhibition of NADP + photoreduction with DCIPH 2 without significantly affecting the rate of NADP + photoreduction with water. The implications of these findings for the differential involvement of Photosystem I and Photosystem II in the photoreduction of NADP + with different electron donors are discussed.

Effect of NADP + on light-induced cytochrome changes in membrane fragments from a blue-green alga

The effect of NADP + on light-induced steady-state redox changes of membrane-bound cytochromes was investigated in membrane fragments prepared from the blue-green algae Nostoc muscorum (Strain 7119) that had high rates of electron transport from water to NADP + and from an artificial electron donor, reduced dichlorophenolindophenol (DCIPH 2) to NADP +. The membrane fragments contained very little phycocyanin and had excellent optical properties for spectrophotometric assays. With DCIPH 2 as the electron donor, NADP + had no effect on the light-induced redox changes of cytochromes: with or without NADP +, 715- or 664-nm illumination resulted mainly in the oxidation of cytochrome f and of other component(s) which may include a c-type cytochrome with an α peak at 549 nm. With 664 nm illumination and water as the electron donor, NADP + had a pronounced effect on the redox state of cytochromes, causing a shift toward oxidation of a component with a peak at 549 nm (possibly a c-type cytochrome), cytochrome f, and particularly cytochrome b 559. Cytochrome b 559 appeared to be a component of the main noncyclic electron transport chain and was photooxidized at physiological temperatures by Photosystem II. This photooxidation was apparent only in the presence of a terminal acceptor (NADP +) for the electron flow from water.

Reactions of a Monospecific Antiserum to Ferredoxin-NADP+ - Reductase with Chloroplast Preparations

Abstract A monospecific antiserum to ferredoxin-NADP+-reductase inhibits the diaphorase activity of soluble ferredoxin-NADP+-reductase from chloroplasts. Two states of the molecular structure of the lamellar system have been observed, one of which is the state described earlier by Berzborn. Stroma-freed chloroplasts in this condition are not agglutinated by the antiserum, although a specific adsorption of antibodies to reductase onto the lamellar system was demonstrated by the Coombs test. However, a second type of chloroplast preparations gives direct agglutination upon addition of the antiserum. Apparently, agglutination in this state is not sterically hindered by neighboring protein structures. This type of chloroplast preparations appears swollen under the light micro­ scope, but exhibits high rates of electron transport. Chloroplasts of three types of tobacco have been used which differ in the morphology of their lamellar systems. The green type contains a normal ratio of grana and intergrana regions whereas the other two types have extended intergrana regions with either only small grana or no partitions at all. Comparison of the maximal degree of inhibition of the NADP+-reduction in chloroplasts from these types of tobacco by the antiserum, leads to the conclusion that ferredoxin-NADP+-reductase is located in the grana and the intergrana regions of the lamellar system, in the outer surface of the thylakoid membrane.

Absence of a common intermediate pool among individual enzyme chains of the energy-conservation pathway in chloroplasts of Euglena gracilis.

Tri-n-butyltin chloride is a specific inhibitor that binds stoicheiometrically and irreversibly with the ATP-synthesizing sites of chloroplasts. Titration of Euglena chloroplasts with tri-n-butyltin chloride shows about six ATP-synthesizing sites per molecule of cytochrome c(552) or per 380 molecules of chlorophyll. This system was used to study the possibility of linkage between individual enzyme chains of the energy-conservation pathway or the possible existence of a common pool of an intermediate in this pathway. The inhibition of ATP synthesis by tri-n-butyltin chloride at low rates of electron transport (low light-intensities, NADP(+) or ferricyanide as electron acceptor) agrees with a kinetic model of two to four ATP-synthesizing sites per energy-conservation chain. At high rates of electron transport, however, the results occasionally agree with a model of 20 or more sites per chain. The results are interpreted as indicating the absence of a common intermediate pool, but the presence of a limited degree of linkage between individual chains. It also indicates the presence of two energy-conservation sites in these chloroplasts.

Effect of Salts and Electron Transport on the Conformation of Isolated Chloroplasts. I. Light-Scattering and Volume Changes

Whole chloroplasts isolated from the leaves of spinach (Spinacia oleracea L.) exhibit 2 types of conformational change during electron transport. Amine-uncoupled chloroplasts swell and atebrin-uncoupled chloroplasts shrink. Chloroplasts uncoupled by carbonylcyanide phenylhydrazones and by treatment with ethylenediamine tetraacetic acid do not change their volumes or light-scattering properties during electron transport. Phosphorylating chloroplasts shrink only slightly.The rate and extent of the conformational change parallel the rate of electron transport; both the decrease in turbidity with methylamine and the increase in turbidity with atebrin are rougly proportional to the Hill reaction rate. Consequently the great volume and light-scattering changes which occur in the presence of these uncouplers can be attributed, in part, to the very high rates of uncoupled electron transport. However, for a given rate of electron transport the atebrin-induced scattering increase is very much greater than the increase observed during photophosphorylation.When uncouplers are combined, the carbonylcyanide phenylhydrazone effect (no change) supercedes both the methylamine effect (swelling) and the atebrin effect (shrinking). The methylamine effect supercedes the atebrin (shrinking) and ethylenediamine tetracetic acid (no change) effects. The atebrin effect supercedes the ethylenediamine tetraacetic acid effect. A similar hierarchy of effects is observed with regard to the rate of the uncoupled electron transport.These light-scattering changes of whole chloroplasts reflect similar changes which occur in very small digitonin particles of chloroplasts. Therefore one must look among chloroplast substructures for the basic mechanism of swelling and shrinking.Many salts (including methylamine hydrochloride) cause the chloroplasts to shrink. This phenomenon is not osmotic since comparable osmolarities of sucrose are without effect. Magnesium chloride and calcium chloride are most effective but all salts tested gave major volume decrease when less than 0.05 m. The salt-shrunken chloroplasts show greater light-scattering changes during electron transport than do low-salt chloroplasts.

The swelling and shrinking of chloroplasts during electron transport in the presence of phosphorylation uncouplers.

Abstract Illuminated chloroplasts undergo light-scattering changes which are associated with volume changes. These changes can be greatly increased in the presence of electron transport and the increase is quantitatively related to the Hill reaction rate. Chloroplasts treated with the uncoupler atebrin (quinacrine) are capable of very high Hill reaction rates and shrink a great deal during electron transport. The shrinking is reversed rapidly if the light is turned off and even more rapidly if the electron acceptor is exhausted. Chloroplasts treated with the uncoupler methylamine are capable of even higher Hill reaction rates but such chloroplasts swell in the light and shrink again in the dark or after the electron acceptor is exhausted. Both the atebrin-stimulated shrinking and the methylamine-stimulated swelling are partially inhibited by the presence of ATP. Carbonylcyanide phenylhydrazones, like atebrin and methylamine, permit high electron-transport rates by uncoupling the Hill reaction from phosphorylation but they completely abolish the conformational changes of chloroplasts.

CALCIUM ION ACCUMULATION AND VOLUME CHANGES OF ISOLATED LIVER MITOCHONDRIA. CALCIUM ION-INDUCED SWELLING.

1. Liver mitochondria suspended in an iso-osmotic buffered potassium chloride medium containing an oxidizable substrate and phosphate accumulated added Ca(2+). During this process H(+) appeared in the medium and the mitochondrial suspension showed increased light-scattering. Respiration was markedly stimulated. 2. The addition of excess of Ca(2+), respiratory inhibitors or uncoupling agents caused extensive mitochondrial swelling associated with release of Ca(2+) into the suspending medium. When the suspension became anaerobic extensive swelling also occurred. Only under conditions when the addition of uncoupling agents would have produced high rates of electron transport, e.g. in the presence of succinate, was the structural integrity of the mitochondrion maintained after Ca(2+) accumulation. 3. Conditions that prevented respiration-dependent Ca(2+) accumulation also prevented Ca(2+)-induced swelling. Bovine plasma albumin was without effect, indicating that U-factor was not involved. Oligomycin together with ADP or ATP partially stabilized the mitochondria against Ca(2+)-induced swelling. 4. It is suggested that a ;high-energy' intermediate generated by coupled electron transport is required to prevent the mitochondrial swelling that results as a consequence of Ca(2+) accumulation.

Optical rotatory dispersion of cytochrome c phospholipid complexes.

PHOSPHOLIPIDS are primary structural materials of the mitochondrial membrane, and numerous observations implicate them in the function of cytochrome c (ref. 1). Thus, when bound in the mitochondrial membrane, cytochrome c is not easily extractable and permits higher rates of electron transport than does added cytochrome c (refs. 2–4). Moreover, when the beef heart mitochondrial electron transport particle is treated with phospholipase, cytochrome c is released from the binding site as the soluble protein5. In addition, phospholipids are required for activity of purified preparations of reduced cytochrome c oxidase6–8, and for reactivation of succinoxidase activity after solvent extraction of mitochondria9.