Effects of 2,5-Dibromo-3-methyl-6-isopropyl-p-benzoquinone (DBMIB) on Light Induced and Glycolytic Phosphate Uptake in Scenedesmus

The effects of DBMIB on photophosphorylation and glycolysis in Scenedesmus obtusiusculus Chod. were investigated by measuring the uptake of inorganic phosphate. To analyze the effects of DBMIB on the different energy coupling possibilities in open chain and cyclic photophosphorylation, DBMIB was given to the algae in narrow concentration intervals between 10^?6M to 10^?4M, either alone, or in combination with DCMU (3-(3,4-dichlorophenyl)-1,1-dimethylurea) or desaspidin. DBMIB inhibits non-cyclic as well as cyclic photophosphorylation in Scenedesmus. However, the DCMU resistant photophosphorylation reactions are less sensitive to DBMIB than the open chain photophosphorylating system in non-DCMU treated cells. Low concentrations of DBMIB even released a part of the DCMU inhibition. Experiments with combinations of DBMIB and desaspidin also indicated that cyclic photophosphorylation is less sensitive to DBMIB than non-cyclic. The inhibition of DCMU resistant cyclic phosphorylation by DBMIB, which is a competitive inhibitor of quinones, indicated a participation of plastoquinones in this type of energy coupling as well as in the non-cyclic and DCMU-sensitive processes. The cyclic and the non-cyclic photophosphorylation pathways probably use different parts of the plastoquinone pool. For the purpose of the experiments, it was necessary to produce data for the effect of DBMIB (10^?6–10^?4M) on glycolysis. The highest concentration gave 50% inhibition.     

Delayed light studies on photosynthetic energy conversion. VIII. Evidence from millisecond emission of chloroplasts for two adenylate binding sites on membrane-bound coupling factor,CF1

Effects of adenylates on chloroplast delayed light emission, at millisecond dark times, are inverse to the previously characterized effects of adenylates on electron transport rates. Either ADP alone or ATP alone increase intensity of delayed light, while ADP plus P_i decrease it. ADP alone requires the presence of an electron acceptor to have this effect on delayed light, but ATP does not.All three adenylate effects are abolished by uncoupling with gramicidin, by partial removal of photophosphorylation coupling factor (CF₁) with EDTA, and by antibody to CF₁. Readdition of CF₁ re-established the adenylate effects in EDTA-stripped membranes. The three adenylate effects are differentially sensitive to pH, and pH differentially affected their abolition by antibody to CF₁. The two adenylate effects shown in the absence of P_i are exhibited at lower adenylate concentrations than the ADP plus P_i effect, and are also less sensitive to phloridzin.These results are discussed in terms of probable adenylate effects on membrane-bound chloroplast coupling factor, CF₁. At least two ADP binding sites would differ with respect to adenylate concentration for half maximal binding; pH of optimal binding capacity; phloridzin sensitivity; and functional regulation of electron transport, proton uptake, and energy storage within the membrane as measured by delayed light emission. It remains unclear whether the high affinity ADP binding site is identical to a high affinity ATP binding site on CF₁.     

Regulation of the cytosolic adenylate ratio as determined by rapid fractionation of mesophyll protoplasts of oat

Adenylate levels in chloroplasts, mitochondria and the cytosol of oat mesophyll protoplasts were determined under light and dark conditions, in the absence and presence of plasmalemma-permeable inhibitors of electron transfer and uncouplers of phosphorylation. This was achieved using a microgradient technique which allowed an integrated homogenization and fractionation of protoplasts within 60 s (Hampp et al. 1982, Plant Physiol. 69, 448–455), under conditions which quench bulk activities of metabolic interconversion in less than 2 s. In illuminated controls, ATP/ADP ratios were found to be 2.1 in chloroplasts, about unity in mitochondria, and 11 in the cytosol; whereas, in the dark, this ratio only showed a large drop in chloroplasts (0.4). None of the compounds used [carbonylcyanide m-chlorophenylhydrazone (CCCP), carbonylcyanide p-trifluoromethoxy-phenylhydrazone (FCCP), antimycin A, dibromothymoquinone (DBMIB), dichlorophenyldi-methylurea (DCMU), or salicylhydroxamic acid (SHAM)] affected the stroma adenylate ratio in the dark. Under illumination, however, the ATP/ADP ratios were partly reduced in the presence of antimycin (inhibitor of cyclic photophosphorylation) and of DCMU (inhibitor of linear electron flow), while in the presence of DBMIB, DCMU+ antimycin (inhibition of both cyclic and linear electron flow), and CCCP (uncoupling) the ratio obtained was the same as that occurring in the dark. In contrast, mitochondrial adenylate levels did not exhibit large variations under the various treatments. The cytosolic ATP/ADP ratio, however, showed dramatic changes: in darkened protoplasts, cytosolic values dropped to 0.2 and 0.1 in the presence of uncouplers and antimycin, respectively, while SHAM did not induce any significant alteration. In the light, a similar pronounced decrease in ATP levels was observed only after the application of uncouplers or inhibitors of both mitochondrial and photosynthetic electron transport, whereas selective inhibition of the latter was largely ineffective in reducing the cytosolic ATP/ADP ratio. Thus, the results show that the antimycin-sensitive electron transport is, potentially, equally active in light and darkness. In addition, they indicate that antimycin-insensitive electron transport in mitochondria (alternative pathway) does not significantly contribute to the cytosolic energy state.Abbreviations CCCP carbonylcyanide m-chlorophenylhydrazone - DBMIB dibromothymoquinone (2,5-dibromo-3-methyl-6-isopropy-p-benzoquinone) - DCMU dichlorophenyldimethylurea - FCCP carbonylcyanide-p-trifluoromethoxy-phenylhydrazone - SHAM sancylhydroxamic acid     

Delayed light studies on photosynthetic energy conversion. VIII. Evidence from millisecond emission of chloroplasts for two adenylate binding sites on membrane-bound coupling factor, CF1.

Effects of adenylates on chloroplast delayed light emission, at millisecond dark times, are inverse to the previously characterized effects of adenylates on electron transport rates. Either ADP alone or ATP alone increase intensity of delayed light, while ADP plus Pi decrease it. ADP alone requires the presence of an electron acceptor to have this effect on delayed light, but ATP does not. All three adenylate effects are abolished by uncoupling with gramicidin, by partial removal of photophosphorylation coupling factor (CF1) with EDTA, and by antibody to CF1. Readdition of CF1 re-established the adenylate effects in EDTA-stripped membranes. The three adenylate effects are differentially sensitive to pH, and pH differentially affected their abolition by antibody to CF1. The two adenylate effects shown in the absence of Pi are exhibited at lower adenylate concentrations than the ADP plus Pi effect, and are also less sensitive to phloridzin. These results are discussed in terms of probable adenylate effects on membrane-bound chloroplast coupling factor, CF1. At least two ADP binding sites would differ with respect to adenylate concentration for half maximal binding; pH of optimal binding capacity; phloridzin sensitivity; and functional regulation of electron transport, proton uptake, and energy storage within the membrane as measured by delayed light emission. It remains unclear whether the high affinity ADP binding site is identical to a high affinity ATP binding site on CF1.     

The Effect of Light Intensity on Total Photophosphorylation and the ATP Level in Scenedesmus

The dependence of in vivo photophosphorylation on light intensity was studied in the unicellular green alga Scenedesmus obtusiusculus. By selective use of the inhibitor DCMU, phosphorylation in (I) the complete system, (II) the pseudocyclic system alone, and (III) the true cyclic system alone, were followed. When the total binding of phosphate was studied, all reaction types became light saturated in about the same manner. The effect of DCMU on the level of ATP varied according to light intensity. As for the specific systems of photophosphorylation, the following ATP data were found: (I) In the complete system the level of ATP decreases with light intensity. (II) Under pseudo-cyclic conditions light first increases and then decreases the ATP level. Under the atmospheric conditions used (i.e. CO₂-free nitrogen) this indicates a regulation between photophosphorylation and glycolysis, for which possible explanations are discussed. (III) In the true cyclic conditions light has little effect on the ATP level. The possibility is indicated that there is a structural difference between the non-cyclic (site 1) and the pseudocyclic (site 2) sites of photophosphorylation on the one hand and the true cyclic site (3) on the other.     

The relationship between state II to state I transitions and cyclic electron flow around photosystem I

The effects of electron acceptors, inhibitors of electron flow and uncouplers and inhibitors of photophosphorylation on a state II to I transition were studied. 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) did not inhibit the state II to I transition. By contrast, 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone (DBMIB), methyl viologen and antimycin A inhibited the transition indicating that the cyclic electron flow around photosystem I, but not the oxidation of electron carriers (such as plastoquinone), induced the state II to I transition. Uncouplers, but not inhibitors of photophosphorylation, inhibited the state transition suggesting that the proton transport through the cyclic electron flow was related to the transition.     

The relationship between state II to state I transitions and cyclic electron flow around photosystem I

The effects of electron acceptors, inhibitors of electron flow and uncouplers and inhibitors of photophosphorylation on a state II to I transition were studied. 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) did not inhibit the state II to I transition. By contrast, 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone (DBMIB), methyl viologen and antimycin A inhibited the transition indicating that the cyclic electron flow around photosystem I, but not the oxidation of electron carriers (such as plastoquinone), induced the state II to I transition. Uncouplers, but not inhibitors of photophosphorylation, inhibited the state transition suggesting that the proton transport through the cyclic electron flow was related to the transition.     

Effects of Light and 3-(3,4-Dichlorophenyl)-1,1-Dimethylurea on Levels of ATP in Lemna paucicostata 6746 and a Photosynthetic Mutant with Abnormal Flowering Responses

The effects of light, 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU), and ammonium ion on pool sizes of ATP were studied in Lemna paucicostata 6746 (wild type) and a photosynthetic mutant (strain 1073) with abnormal flowering responses. Wild type fronds were capable of endogenous and phenazine methosulfate-catalyzed cyclic photophosphorylation. The endogenous cyclic photophosphorylation was inhibited by DCMU. The mutant fronds showed little endogenous but appreciable rates of phenazine methosulfate-catalyzed cyclic photophosphorylation. Treatment with DCMU during prolonged exposure to light did not result in elevated levels of ATP. Ammonium ion in the medium did not inhibit light-induced increases in pool sizes of ATP. It is concluded that the previously reported effects on flowering of DCMU, the photosynthetic mutation or ammonium ion, were not due to altered pool sizes of ATP.     

Phosphorus incorporation in the ehrlich ascites tumor cell

Data from isotopic uptake experiments were used to measure the kinetics of labelling of cellular phosphate, ATP and ADP in the Ehrlich ascites tumor cell. The results show that steady state phosphate exchange flux was 0.333 ± 0.052 (S.E.) μmoles per 10⁷ cells per hour at 37°, and that the specific activity of phosphate was the same as P_γ ATP. Metabolic inhibition reduced the phosphate flux by 30–50%. A model, based on oxidative phosphorylation and the adenylate kinase reaction is used to interpret the labelling sequence of P_β ATP and P_β ADP, and its dependence on P_γ ATP.     

Inhibition of photophosphorylation by ATP and the role of magnesium in photophosphorylation

ATP and pyrophosphate at high concentration (> 1 mM) inhibited photophosphorylation of isolated spinach chloroplasts in the normal salt medium and did not cause stimulation of electron transport. The inhibition of photophosphorylation by ATP or pyrophosphate was shown to be abolished by the addition of excess MgCl₂, ADP and phosphate. It has been demonstrated that the rates of photophosphorylation in the absence and presence of ATP or pyrophosphate are determined similarly by the concentrations of magnesium-ADP (Mg · ADP^?) and magnesium-phosphate (Mg · P_i) complexes.It is highly probable that Mg · ADP^? and Mg · P_i, but not free ADP and free phosphate, are the active form of the substrates of photophosphorylation. This is in support of the view that ATP inhibits photophosphorylation by decreasing the concentration of Mg²⁺ which is available for the formation of the complex with ADP and phosphate.     

The ATP Levels Obtained in Photophosphorylation, Glycolysis and Oxidative Phosphorylation in Scenedesmus Titrated with Desaspidin

The ATP levels in photophosphorylation, glycolysis and oxidative phosphorylation, in the unicellular green alga Scenedesmus obtusiusculus, were titrated with narrow concentration intervals of desaspidin in the presence of different concentrations of 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU), which allows the differentiation between non-cyclic, pseudocyclic and true cyclic photophosphorylation. The data on photophosphorylative ATP levels were compared with earlier data on total binding of phosphate. In the true cyclic process, both parameters are equally sensitive towards desaspidin. Under pseudocyclic conditions and in non-cyclic photophosphorylation, the level of ATP is more sensitive towards desaspidin than is total binding of phosphate. This suggests a structural difference between the cyclic and the two non-cyclic (one of which is also pseudocyclic) sites. The non-cyclic ATP level is more sensitive towards desaspidin than is pseudocyclic. This may be connected with the higher ATP level under pseudocyclic as compared to non-cyclic conditions.     

ATP pools and transients in the blue-green alga,Anabaena cylindrica

Anabaena cylindrica grown in steady state continuous culture has an extractable ATP pool, measured on the basis of the luciferin-luciferase assay of 165±35 nmoles ATP mg chla ^-1. This pool is maintained by a dynamic balance between the rate of ATP synthesis and the rate of ATP utilization. Phosphorylating mechanisms which can maintain the pool in the short term are total photophosphorylation, cyclic photophosphorylation and oxidative phosphorylation. The alga can maintain its ATP pool by switching rapidly from one of these phosphorylating mechanisms to another depending on the environmental conditions. At each switch-over there is a transient drop in the ATP pool for a few seconds. On switching to conditions where only substrate level phosphorylation operates, the ATP pool falls immediately, but takes several hours to recover. The apparent rates of ATP synthesis by total photophosphorylation and by cyclic photophosphorylation are both much higher (210±30 and 250±13 moles ATP mg chla ^-1 h^-1 respectively) than the apparent rate of ATP synthesis by oxidative phosphorylation (22±3 moles ATP mg chla ^-1 h^-1). In long term experiments the ATP pool is maintained when total photophosphorylation is operating. It cannot be maintained in the long term by cyclic photophosphorylation alone in the absence of photosystem II activity or endogenous carbon compounds, or by oxidative phosphorylation in the absence of endogenous carbon compounds. Measurements of ATP, ADP and AMP show that the total pool of adenylates is similar in the light and in the dark in the short term. There is only limited production of ATP under dark anaerobic conditions when glycolysis and substrate phosphorylation can operate which suggests that these processes are of limited significance in providing ATP in Anabaena cylindrica.Abbreviations ADP adenosine 5-diphosphate - AMP adenosine 5-monophosphate - ATP adenosine 5-triphosphate - CCCP carbonyl cyanide m-chlorophenyl hydrazone - DCMU 3-(3,4-dichlorophenyl)1,1-dimethyl urea - HEPES N-2-hydroxyethylpiperazine-N-2-ethane sulfonic acid - PEP phosphoenolpyruvate     

Photophosphorylation in Scenedesmus in vivo: O2 evolution, ATP pools and transients, and phosphate binding during photoreduction of NO3−, NO2− and CO2

The relation between light-induced electron transport with NO₃^?, NO₂^? or CO₂ as acceptors, ATP pools and transients in dark-light-dark transitions, and phosphate uptake was examined in phosphorus-starved cells of Scenedesmus obtusiusculus Chod. Net O₂ evolution at saturating light was around 6 μmol × (mg chlorophyll × h)^?1 in the absence of any acceptor, but reached average rates of 21, 65 and 145 μmol × (mg chlorophyll × h)^?1 upon additions of 5 mM KNO₃, KNO₂ and KHCO₃, respectively. The apparent rate of photophosphorylation in transition experiments was only a few percent of the rate calculated from CO₂-dependent O₂ evolution. Blocking non-cyclic electron transport with DCMU inhibited phosphate assimilation, but acceleration of non-cyclic electron flow by addition of NO₃^? or NO₂^? did not stimulate phosphate assimilation as compared to the situation without an acceptor. A functional non-cyclic system might primarily be needed for an efficient shuttle transfer of ATP from the chloroplast to the cytoplasm. An inhibition of the non-cyclic system due to lack of reducible substrates accelerates the cyclic system and thus indicates a regulation mechanism between the two systems.     

Effects of various buffers, 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) and NaN3 on menadione mediated photophosphorylation in isolated chloroplasts

The effects of DCMU and NaN₃ were studied on menadione-mediated photophosphorylation in broken spinach chloroplasts kept in low oxygen tension in Tricine or HEPES buffers at either high or reduced irradiances. – (A) At high irradiance (131 W. m^?2) and absence of NaN₃ the ATP formation was inhibited by DCMU regardless of the type of buffer used. – (B) At high irradiance and presence of NaN₃ some concentrations of DCMU stimulated, whilst others inhibited the ATP formation in a HEPES buffer. The ATP formation was predominantly inhibited by DCMU in a Tricine buffer. – (C) At reduced irradiance (57 W. m^?2) the chloroplasts in a HEPES buffer were almost insensitive towards DCMU both in the presence and absence of NaN₃. – (D) Chloroplasts in a Tricine buffer were slightly stimulated in their ATP formation by DCMU at reduced irradiance either with or without the presence of NaN₃ in the experimental medium. When menadione acts as a terminal electron acceptor, oxygen is consumed on its reoxidation. The results indicate that this process may occur with oxygen released by the splitting of water as the main oxidant. – The data also demonstrate the importance of caution when selecting buffering substances as well as when choosing light intensities for experiments on photophosphorylation in chloroplasts.     

Fatty acid synthesis by spinach chloroplasts III. Relationship between fatty acid synthesis and photophosphorylation

The modes of actions of photosynthetic inhibitors on photosynthesisand fatty acid synthesis were examined. DCMU, an electron transport inhibitor, inhibited fatty acidsynthesis and photophosphorylation to the same extent, suggestingdependence of fatty acid synthesis on photosynthesis. The samewas also the case with FCCP, a photophosphorylation uncoupler.In contrast, NH₄Cl and phlorizin at concentrations completelysuppressing ATP formation, only partially inhibited the fattyacid synthesis. These facts suggest that a certain level ofhigh-energy intermediate (state) is responsible for the lightenhancement of fatty acid synthesis. This idea is further supportedby the fact that the partial inhibition of fatty acid synthesisby NH₄Cl was relieved by addition of DCCD at low concentrationssuppressing the ATP formation but not completely destroyingthe high energy intermediate. The lag period in the initial period of fatty acid synthesiswas shortened by preillumination of chloroplasts, even in theabsence of ADP. This indicates that the light dependent fattyacid synthesis is closely associated with the high-energy intermediate(state), but not directly with ATP formation by photophosphorylation. ¹ Present address: Radioisotope Centre, University of Tokyo,Yayoi, Bunkyo, Tokyo 113, Japan. (Received August 26, 1974; )     

Titration of Photophosphorylation in Spinach Chloroplasts with 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU)

The kinetics of the inhibition of photophosphorylation in chloroplasts from spinach (Spinacia oleracea) was investigated with 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) in small concentration intervals, starting at 10^-7M. Plots of the reciprocal of photophosphorylation against concentration of DCMU gave essentially the same straight line with 2 mM nicotinamide adenine dinucleotide phosphate (NADP) together with saturating amounts of ferredoxin or with 4 mM K₃Fe(CN)₆ as the final acceptors for electrons. Practically complete inhibition was obtained at 3 x 10^-6M DCMU. With 0.1 mM flavin mononucleotide (FMN) and ferredoxin, the inhibition between 10^-7M and 10^-6M DCMU was a little slower than in the other two cases. At 10^-6M DCMU a break occurred to a new straight line in the plots, indicating that another reaction was inhibited. Total photophosphorylation without DCMU was about 77 μmol ATP per mg chlorophyll and hour. At the breaking point 20% remained, and inhibition was not complete even at 8 x 10^-6M DCMU. The inhibitor constant for the high-DCMU reaction was in the order of 2 x 10^-5M; for the low-DCMU reaction some complication made the “constant” appear negative. With phenazine methosulfate (PMS) added, DCMU was without effect on photophosphorylation. – As earlier shown by us, titration curves for intact cells of the microalga Scenedesmus show the break at 10^-6M DCMU; and above 6 x 10^-6M photophosphorylation in the algae is not further decreased by DCMU. The data are compared and their possible significance is discussed.     

Studies on the pathway of cyclic electron flow in mesophyll chloroplasts of a C4 plant

1. Cyclic photophosphorylation driven by white light, as followed by ¹⁴CO₂ fixation by mesophyll chloroplast preparations of the C₄ plant Digitaria sanguinalis, was specifically inhibited by disalicylidenepropanediamine (DSPD), antimycin A, 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone (DBMIb), 1-ethyl-3(3-dimethylaminopropyl)-carbodiimide (EDAC), and KCN suggesting that ferredoxin, cytochrome b₅₆₃, plastoquinone, cytochrome f, and plastocyanin are obligatory intermediates of cyclic electron flow. It was found that 0.2 μM DCMU and 40 μM o-phenanthroline blocked noncyclic electron flow, stimulated cyclic photophosphorylation, and caused a partial reversal (40–100%) of the inhibition by DBMIB and antimycin A, but not DSPD.

2. Cyclic photophosphorylation could also be activated using only far-red illumination. Under this condition, however, cyclic photophosphorylation was much less sensitive to the inhibitors DBMIB, EDAC and antimycin A, but remained completely sensitive to DSPD and KCN. Inhibition in far-red light was not increased by preincubating the chloroplasts with the various inhibitors for several minutes in white light.

3. The striking correspondence between the effects of photosystem II inhibitors, DCMU and o-phenanthroline, on cyclic photophosphorylation under white light and cyclic photophosphorylation under far-red light (in the absence of photosystem II inhibitors) suggests that electrons flowing from photosystem II may regulate the pathway of cyclic electron flow.     

Synergistic uncoupling of spinach chloroplasts by combinations of photophosphorylation inhibitors and carbonyl cyanide p-trifluoromethoxyphenylhydrazone.

Combinations of low concentrations of carbonyl cyanide p-trifluoromethoxyphenylhy-drazone (FCCP) with suboptimal concentrations of Dio-9, phloridzin, ajmaline, and dihydrodiscarine B synergistically inhibited cyclic and noncyclic photophosphorylation in spinach chloroplasts but their effects on the light-triggered ATPase were additive rather than synergistic. The effect was reversed by washing and prevented by dithioerythritol and by cistein. Carbonyl cyanide m-chlorophenylhydrazone (CCP) could replace FCCP but uncouplers of other types like atebrin did not substitute for FCCP.Combinations of FCCP with the four inhibitors synergistically uncoupled ferricyanide reduction in the presence of ADP and P_i but not in their absence. The synergistic uncoupling was not observed on the light-dependent pH rise of chloroplast suspensions.Association of FCCP with any of the inhibitors completely abolished the stimulation of proton uptake or the inhibition of electron transport induced by low concentrations of ATP.This synergistic and peculiar uncoupling can not be ascribed to a modification of membrane permeability. One possible explanation is that the effect requires a conformational state of the membrane-bound coupling factor 1 (CF₁) induced by phosphorylating conditions which would facilitate the interaction of inhibitors and FCCP with the membrane.     

Anaerobic ATP Levels in the Blue-Green Alga Anabaena: Dark-Light Transients and Effects of Light Intensity

An enzymatic method was used to determine ATP extracted from anaerobically incubated samples of the blue-green alga Anabaena variabilis. The dark-light-dark transients of the ATP level in the whole cells were studied in the presence and in the absence of 3 (p-chlorophenyl)-l.l-dimethylurea (CMU). When O₂ evolution was completely inhibited by CMU, there was no significant difference between the transient under illumination of a low intensity and that of fourfold intensity. On the other hand in the absence of CMU, the ATP level decreased more rapidly after high intensity illumination than after low intensity one. The increase in ATP content during the first 5 s illumination was smaller in the absence of CMU than in the presence of CMU. With or without CMU the increase was saturated at the vicinity of the intensity at which photosynthetic O₂ evolution compensates for respiratory O₂ consumption. The calculated quantum requirement of cyclic photophosphorylation in vivo was 2.2 quanta per ATP.     

Sulphate uptake in the unicellular marine red algaRhodella maculata

Sulphate uptake by the unicellular marine red algaRhodella maculata conforms to Michaelis-Menten kinetics. Two uptake systems have been found: a low affinity system with an apparentK _m of 22 mM, and a high affinity system with an apparentK _m of 63.4 M. Transition from the low to the high affinity system can occur within 2.5 min, in response to a decrease in the ambient sulphate concentration to below 10 mM. Assimilation rates in the dark are about 20% those in the light, although enhancement by light is independent of the quanlity of light supplied above 27 mol m^-2 s^-1. Use of metabolic inhibitors indicates that photophosphorylation provides the main source of energy for sulphate assimilation, through both cyclic and non-cyclic electron flow.Abbreviations used APS-kinase ATP:adenylyl-sulphate 3-phosphotransferase (E.C. 2.7.1.25) - ATP-sulphurylase ATP:sulphate adenylyltransferase (E.C.2.7.74) - DCMU [3-(3,4-dichlorophenyl)]-1,1 dimethylurea - 2,4 DNP 2,4-dinitrophenol - DBMIB Dibromothymoquinone (2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone)