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1.
Driving Forces for Bicarbonate Transport in the Cyanobacterium Synechococcus R-2 (PCC 7942) 总被引:4,自引:0,他引:4
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Air-grown Synechococcus R-2 (PCC 7942) cultures grown in BG-11 medium are very alkaline (outside pH is 10.0) and use HCO3- as their inorganic carbon source. The cells showed a dependence on Na+ for photosynthesis, but low Na+ conditions (1 mol m-3) were sufficient to support saturating photosynthesis. The intracellular dissolved inorganic carbon in the light was greater than 20 mol m-3 in both low-Na+ conditions and in BG-11 medium containing the usual [Na+] (24 mol m-3, designated high-Na+ conditions). The electrochemical potential for HCO3- in the light was in excess of 25 kJ mol-1, even in high-Na+ conditions. The Na+-motive force was greater than -12 kJ mol-1 under both Na+ conditions. On thermodynamic grounds, an Na+-driven co-port process would need to have a stoichiometry of 2 or greater ([greater than or equal to]2Na+ in/HCO3-1 in), but we show that Na+ or K+ fluxes cannot be linked to HCO3- transport. Na+ and K+ fluxes were unaffected by the presence or absence of dissolved inorganic carbon. In low-Na+ conditions, Na+ fluxes are too low to support the observed net 14C-carbon fixation rate. Active transport of HCO3- hyperpolarizes (not depolarizes) the membrane potential. 相似文献
2.
Uptake of Thallium, a Toxic Heavy-Metal, in the Cyanobacterium Synechococcus R-2 (Anacystis nidulans, S. Leopoliensis) PCC 7942 总被引:1,自引:0,他引:1
Uptake of the toxic heavy-metal, thallium, was studied in thecyanobacterium Synechococcus R-2 (PCC 7942) using clinicallyavailable 201Tl +. Thallium was found to distribute across theplasmalemma passively, and so the accumulation ratio of theion ([Tl+]i/[Tl+]o) could be used to calculate the apparentmembrane potential (i,o) of the cells (ETI+i,o = i,o).The permeability of the plasmalemma to TI+ (PTI+ 1 to 5 nms1)is higher than that of K+. Valinomycin does not increase thepermeability of TI+. Transient changes in the i,o of cells,because of electrogenic transport of ions, could be detectedfrom its effects upon the uptake rate of TI+. HCO3 hyperpolarizedSynechococcus cells, whereas NH+4, CH3NH+, and K+ led to depolarization.The use of TI+ as a reporter of i,o has some inherent limitations.Tl+ is toxic at very low concentrations (inhibitory effectsare apparent after about 6 h at concentrations as low as 1 mmolm3). The rate of equilibration is slow (t1/25 to 20 min).Equilibration of TI+ takes about 2 h, which limits its valueas a membrane potential probe. Large amounts of TI+ bind tothe surface of the cells making the method impracticable formeasuring accumulation ratios of less than about 10 (i,o)values smaller than about 60 mV). Cultures continuouslyexposed to Tl+ (10 mmol m3) eventually become TI+ resistantby actively extruding TI+ (µTI+i,o= 3±0.2kJ mol1) and so thallium cannot be used as a i,oprobe in such cells. (Received October 28, 1997; Accepted August 31, 1998) 相似文献
3.
Carbonic Anhydrase Activity Associated with the Cyanobacterium Synechococcus PCC7942 总被引:12,自引:5,他引:7
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Intact cells and crude homogenates of high (1% CO2) and low dissolved inorganic carbon (Ci) (30-50 microliters per liter of CO2) grown Synechococcus PCC7942 have carbonic anhydrase (CA)-like activity, which enables them to catalyze the exchange of 18O from CO2 to H2O. This activity was studied using a mass spectrometer coupled to a cuvette with a membrane inlet system. Intact high and low Ci cells were found to contain CA activity, separated from the medium by a membrane which is preferentially permeable to CO2. This activity is most apparent in the light, where 18O-labeled CO2 species are being taken up by the cells but the effluxing CO2 has lost most of its label to water. In the dark, low Ci cells catalyze the depletion of the 18O enrichment of CO2 and this activity is inhibited by both ethoxyzolamide and 2-(trifluoromethoxy)carbonyl cyanide. This may occur via a common inhibition of the Ci pump and the Ci pump is proposed as a potential site for the exchange of 18O. CA activity was measurable in homogenates of both cell types but was 5- to 10-fold higher in low Ci cells. This was inhibited by ethoxyzolamide with an I50 of 50 to 100 micromolar in both low and high Ci cells. A large proportion of the internal CA activity appears to be pelletable in nature. This pelletability is increased by the presence of Mg2+ in a manner similar to that of ribulose bisphosphate carboxylase-oxygenase activity and chlorophyll (thylakoids) and may be the result of nonspecific aggregation. Separation of crude homogenates on sucrose gradients is consistent with the notion that CA and ribulose bisphosphate carboxylase-oxygenase activity may be associated with the same pelletable fraction. However, we cannot unequivocally establish that CA is located within the carboxysome. The sucrose gradients show the presence of separate soluble and pelletable CA activity. This may be due to the presence of separate forms of the enzyme or may arise from the same pelletable association which is unstable during extraction. 相似文献
4.
Purification and Characterization of Phosphoribulokinase from the Cyanobacterium Synechococcus PCC7942 总被引:1,自引:0,他引:1
Wadano Akira; Kamata Yoichi; Iwaki Toshio; Nishikawa Keisuke; Hirahashi Tomohiro 《Plant & cell physiology》1995,36(7):1381-1385
Phosphoribulokinase (PRK) was purified to electrophoretic homogeneityfrom Synechococcus PCC7942 with high specific activity. Molecularmasses of the native enzyme and its subunit were 178 and 42kDa, respectively. Cys-17 and Cys-38 were conserved in the cyanobacterialPRK, but 18 amino acid residues between them were missing amongthe 40 residues found in higher plant PRKs. (Received February 1, 1995; Accepted July 27, 1995) 相似文献
5.
Ethoxyzolamide Inhibition of CO(2)-Dependent Photosynthesis in the Cyanobacterium Synechococcus PCC7942 总被引:5,自引:4,他引:1
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Cells of the cyanobacterium, Synechococcus PCC7942, grown under high inorganic carbon (Ci) conditions (1% CO2; pH 8) were found to be photosynthetically dependent on exogenous CO2. This was judged by the fact that they had a similar photosynthetic affinity for CO2 (K0.5[CO2] of 3.4-5.4 micromolar) over the pH range 7 to 9 and that the low photosynthetic affinity for Ci measured in dense cell suspensions was improved by the addition of exogenous carbonic anhydrase (CA). The CA inhibitor, ethoxyzolamide (EZ), was shown to reduce photosynthetic affinity for CO2 in high Ci cells. The addition of 200 micromolar EZ to high Ci cells increased K0.5(CO2) from 4.6 micromolar to more than 155 micromolar at pH 8.0, whereas low Ci cells (grown at 30 microliters CO2 per liter of air) were less sensitive to EZ. EZ inhibition in high and low Ci cells was largely relieved by increasing exogenous Ci up to 100 millimolar. Lipid soluble CA inhibitors such as EZ and chlorazolamide were shown to be the most effective inhibitors of CO2 usage, whereas water soluble CA inhibitors such as methazolamide and acetazolamide had little or no effect. EZ was found to cause a small drop in photosystem II activity, but this level of inhibition was not sufficient to explain the large effect that EZ had on CO2 usage. High Ci cells of Anabaena variabilis M3 and Synechocystis PCC6803 were also found to be sensitive to 200 micromolar EZ. We discuss the possibility that the inhibitory effect of EZ on CO2 usage in high Ci cells of Synechococcus PCC7942 may be due to inhibition of a `CA-like' function associated with the CO2 utilizing Ci pump or due to inhibition of an internal CA activity, thus affecting CO2 supply to ribulose bisphosphate carboxylase-oxygenase. 相似文献
6.
Ascorbate peroxidase active component (APAC) was purified and characterized in Synechococcus PCC 9742 (R2) cells. APAC was isolated from freshly harvested cells, by ion exchange chromatography on DEAE cellulose, ultrafiltration through a 3000 dalton cut off filter and high pressure liquid chromatography through a reversed phase C-18 column. APAC was found to be extremely stable to harsh treatments of boiling water for 30 min, acidification to pH 2.0 and proteolytic digestion. A close correlation between activity and iron content of APAC was observed throughout the purification steps. E.S.R. spectrum of APAC showed a resonance line at g = 4.3 in the oxidized from. Peroxide reduction by ascorbate decreased the E.S.R. signal, which reappeared upon reoxidation by H2O2. The affinities of APAC to H2O2 and ascorbate were high (0.38 mM and 0.2 mM, respectively). Amino acid composition analysis of APAC revealed the presence of glutamic acid: glycine: cysteine residues at 2: 1: 1 ratio. 相似文献
7.
Iron Superoxide Dismutase Protects against Chilling Damage in the
Cyanobacterium Synechococcus species
PCC7942 总被引:1,自引:0,他引:1
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David J. Thomas Jannette B. Thomas Shane D. Prier Nicole E. Nasso Stephen K. Herbert 《Plant physiology》1999,120(1):275-282
A strain of Synechococcus sp. PCC7942 lacking functional Fe superoxide dismutase (SOD), designated sodB−, was characterized by its growth rate, photosynthetic pigments, inhibition of photosynthetic electron transport activity, and total SOD activity at 0°C, 10°C, 17°C, and 27°C in moderate light. At 27°C, the sodB− and wild-type strains had similar growth rates, chlorophyll and carotenoid contents, and cyclic photosynthetic electron transport activity. The sodB− strain was more sensitive to chilling stress at 17°C than the wild type, indicating a role for FeSOD in protection against photooxidative damage during moderate chilling in light. However, both the wild-type and sodB− strains exhibited similar chilling damage at 0°C and 10°C, indicating that the FeSOD does not provide protection against severe chilling stress in light. Total SOD activity was lower in the sodB− strain than in the wild type at 17°C and 27°C. Total SOD activity decreased with decreasing temperature in both strains but more so in the wild type. Total SOD activity was equal in the two strains when assayed at 0°C. 相似文献
8.
Nitrite-Specific Active Transport System of the Cyanobacterium Synechococcus sp. Strain PCC 7942 总被引:1,自引:0,他引:1
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Shin-ichi Maeda Masato Okamura Masaki Kobayashi Tatsuo Omata 《Journal of bacteriology》1998,180(24):6761-6763
Studies on the nitrite uptake capability of a mutant of Synechococcus sp. strain PCC 7942 lacking the ATP-binding cassette-type nitrate-nitrite-bispecific transporter revealed the occurrence of a nitrite-specific active transport system with an apparent Km (NO2−) of about 20 μM. Similar to the nitrate-nitrite-bispecific transporter, the nitrite-specific transporter was reversibly inhibited by ammonium in the medium. 相似文献
9.
Ethoxyzolamide Inhibition of CO(2) Uptake in the Cyanobacterium Synechococcus PCC7942 without Apparent Inhibition of Internal Carbonic Anhydrase Activity 总被引:8,自引:6,他引:2
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In high inorganic carbon grown (1% CO2 [volume/volume]) cells of the cyanobacterium Synechococcus PCC7942, the carbonic anhydrase (CA) inhibitor, ethoxyzolamide (EZ), was found to inhibit the rate of CO2 uptake and to reduce the final internal inorganic carbon (Ci) pool size reached. The relationship between CO2 fixation rate and internal Ci concentration in high Ci grown cells was little affected by EZ. This suggests that in intact cells internal CA activity was unaffected by EZ. High Ci grown cells readily took up CO2 but had little or no capacity for HCO3− uptake. These cells appear to possess a CO2 utilizing Ci pump that has a CA-like function associated with the transport step such that HCO3− is the species delivered to the cell interior. This CA-like step may be the site of inhibition by EZ. Low Ci grown cells possess both CO2 uptake and HCO3− uptake activities and EZ inhibited both activities to a similar degree, suggesting that a common step in CO2 and HCO3− uptake (such as the Ci pump) may have been affected. The inhibitor had no apparent effect on internal CO2/HCO3− equilibria (internal CA function) in low Ci grown cells. 相似文献
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11.
Anabaena PCC 7119 showed higher rates of phosphate uptake whencells were under P-starvation. Phosphate uptake was energy-dependentas indicated the decrease observed when assays were performedin the dark or in the presence of inhibitors of photosyntheticelectron transport, energy transfer and adenosine triphosphataseactivity. Phosphate uptake was stimulated by Na+ both in P-sufficientcells and P-starved cells. Li+ and K+ acted as partial analoguesfor Na+. The Na+-stimulation of phosphate uptake followed Michaelis-Mentenkinetics, half-saturation (K) of phosphate uptake was reachedwith a Na+ concentration of 212 µM. The absence of Na+reduced the rates of phosphate uptake at all phosphate concentrationsassayed (120 µM). The maximum uptake rates (Vmax)decreased from 658 nmol P (mg dry wt)-1 h-1 in the presenceof Na+ to 149 nmol P (mg dry wt)-1 h-1 in the absence of Na+.The absence of Na+ did not change significantly the concentrationof phosphate required to reach half-saturation (K) (3.01 µMin the presence of Na+ vs 3.21 µM in the absence of Na+).In the presence of Na+ the rate of phosphate uptake was affectedby the pH; optimal rates were observed at pH 8. In the absenceof Na+ phosphate uptake was not affected by the pH; low rateswere observed in all cases. Monensin, an ionophore which collapsesNa+-gradients, reduced the rate of phosphate uptake in Na+-supplementedcells. These results indicated the existence of a Na+-dependentphosphate uptake in Anabaena PCC 7119. (Received September 8, 1992; Accepted November 17, 1992) 相似文献
12.
Carbon Oxysulfide Is an Inhibitor of Both CO(2) and HCO(3) Uptake in the Cyanobacterium Synechococcus PCC7942 总被引:2,自引:2,他引:0
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Carbon oxysulfide (COS) was reinvestigated as an inhibitor of active inorganic carbon transport in cells of Synechococcus PCC7942 adapted to growth at low inorganic carbon. COS inhibited both CO2 and HCO3− transport processes in a reversible (in the short term) and mixed competitive manner. The inhibition of COS was established using both silicone oil centrifugation experiments and O2-evolution studies. The Ki for COS inhibition was 29 micromolar for CO2 transport and 110 micromolar for HCO3− transport. These results support a model of inorganic carbon transport with a central CO2 pump and an inducible HCO3− utilizing accessory protein which supplies CO2 to the primary pump. 相似文献
13.
Characterization of the dnaK multigene family in the Cyanobacterium Synechococcus sp. strain PCC7942
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The cyanobacterium Synechococcus sp. strain PCC7942 has three dnaK homologues (dnaK1, dnaK2, and dnaK3), and a gene disruption experiment was carried out for each dnaK gene by inserting an antibiotic resistance marker. Our findings revealed that DnaK1 was not essential for normal growth, whereas DnaK2 and DnaK3 were essential. We also examined the effect of heat shock on the levels of these three DnaK and GroEL proteins and found a varied response to heat shock, with levels depending on each protein. The DnaK2 and GroEL proteins exhibited a typical heat shock response, that is, their synthesis increased upon temperature upshift. In contrast, the synthesis of DnaK1 and DnaK3 did not respond to heat shock; in fact, the level of DnaK1 protein decreased. We also analyzed the effect of overproduction of each DnaK protein in Escherichia coli cells using an inducible expression system. Overproduction of DnaK1 or DnaK2 resulted in defects in cell septation and formation of cell filaments. On the other hand, overproduction of DnaK3 did not result in filamentous cells; rather a swollen and twisted cell morphology was observed. When expressed in an E. coli dnaK756 mutant, dnaK2 could suppress the growth deficiency at the nonpermissive temperature, while dnaK1 and dnaK3 could not suppress this phenotype. On the contrary, overproduction of DnaK1 or DnaK3 resulted in growth inhibition at the permissive temperature. These results suggest that different types of Hsp70 in the same cellular compartment have specific functions in the cell. 相似文献
14.
Regulation of Nitrite Reductase Activity under CO2 Limitation in the Cyanobacterium Synechococcus sp. PCC7942 总被引:2,自引:0,他引:2
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During photoautotrophic growth under CO2-limited conditions, cells of Synechococcus sp. PCC7942 excreted into the medium about 30% of the nitrite produced by reduction of nitrate. No nitrite was excreted under CO2-sufficient conditions. After transfer of high-CO2-grown cells to CO2-limited conditions, nitrite reductase activity started to decline within 0.5 h and decreased to 50% of the initial level in 3 h, whereas nitrate reductase activity was virtually unchanged. Nitrite started to accumulate in the medium about 3 h after the transfer of the cells to CO2-limited conditions and reached a concentration of >0.4 mM at 17 h. These findings suggested that the nitrite excretion was due to an imbalance of the activities of nitrite reductase and nitrate reductase. Since ammonium, the product of nitrite reduction, was not detected in the medium, it was concluded that the step of nitrite reduction limits the rate of nitrate assimilation under CO2-limited conditions. The extent of decrease in nitrite reductase activity under CO2-limited conditions was much larger than that caused by rifampicin (an inhibitor of RNA synthesis) treatment under high-CO2 conditions. Addition of CO2, in the form of sodium bicarbonate, to the CO2-limited culture increased the nitrite reductase activity, but rifampicin inhibited this increase. These findings suggested the presence of a mechanism that irreversibly inactivates nitrite reductase under CO2-limited conditions. 相似文献
15.
Association of Carbonic Anhydrase Activity with Carboxysomes Isolated from the Cyanobacterium Synechococcus PCC7942 总被引:1,自引:1,他引:0
The development of a simple method for the isolation of purified carboxysomes from the cyanobacterium Synechococcus PCC7942 has made it possible to identify a specific and inducible, intracellular carbonic anhydrase (CA) activity that is strongly associated with carboxysomes. This was shown, in part, through enzyme recovery experiments that indicated that a clear majority of a CA activity that is sensitive to the CA inhibitor ethoxyzolamide (I50 = 4 μm) copurifies with a majority of the cell's ribulose-1,5-bisphosphate carboxylase/oxygenase activity in a highly purified pelletable fraction. Electron microscopy of this pelletable fraction revealed the presence of carboxysomes that were physically intact. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of carboxysome proteins showed that the large and small subunits of ribulose-1,5-bisphosphate carbosylase/oxygenase were clearly prominent and that several other minor proteins could be distinguished. The specific location of this carboxysomal CA activity is further reinforced by the finding that a previously isolated high CO2-requiring mutant, Type II/No. 68 (G.D. Price, M.R. Badger [1989] Plant Physiol 91: 514-525), displayed a 30-fold reduction in carboxysome-associated CA activity when tested under optimal conditions. Carboxysomal CA has the unusual property of being inactivated by dithiothreitol. The enzyme also requires 20 mm Mg2+ (as MgSO4) for near maximum activity; other divalent cations, such as Ca2+ and Mn2+, also stimulate carboxysomal CA activity, but to a lesser extent than Mg2+. Results are discussed in relation to the role of carboxysomes in the CO2-concentrating mechanism in cyanobacteria and the role that carboxysomal CA activity appears to play in this process. 相似文献
16.
Isolation of a Putative Carboxysomal Carbonic Anhydrase Gene from the Cyanobacterium Synechococcus PCC7942 总被引:6,自引:1,他引:5
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The Type II mutants of the cyanobacterium Synechococcus PCC7942 (G.D. Price, M.R. Badger [1989] Plant Physiol 91: 514-525) are able to accumulate a large pool of inorganic carbon inside the cell, but are unable to utilize it for CO2 fixation, resulting in a high CO2-requiring phenotype. We have isolated a 3.5-kb BamHI clone (pT2) that complements the Type II mutants, and complementation analysis with DNA subclones indicated that the complementing region was located in the 0.75-kb XhoI-Bg/II fragment. This same region hybridized to the chloroplastic carbonic anhydrase (CA) gene from spinach on Southern blots and to a mRNA of approximate 1 kb on northern blots. Restriction mapping and sequence analysis revealed that pT2 is the same as a genomic clone (pBM3.8) that complements another high CO2-requiring (temperature sensitive) mutant, C3P-O (E. Suzuki, H. Fukuzawa, S. Miyachi [1991] Mol Gen Genet 226: 401-408). Recently, a 272-amino acid open reading frame showing 22% homology with pea and spinach chloroplast CA genes was identified in clone pBM3.8 (H. Fukuzawa, E. Suzuki, Y. Komukal, S. Miyachi [1992] Proc Natl Acad Sci USA 89: 4437-4441). CA activity was detected in Escherichia coli cells transformed with subclones of pT2 (pT2-A and pT2-A1) containing the HindIII-Bg/II fragment, and the expressed CA has properties similar to those of the CA activity associated with carboxysomes purified from Synechococcus PCC7942 (G.D. Price, J.R. Coleman, M.R. Badger [1992] Plant Physiol 100: 784-793). Therefore, it is reasonable to conclude that the HindIII-Bg/II fragment codes for the carboxysomal CA gene product. The result is discussed in the context of the role that carboxysomal CA plays in the operation of the CO2-concentrating mechanism in cyanobacteria. 相似文献
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《Bioscience, biotechnology, and biochemistry》2013,77(5):1394-1397
Uroporphyrinogen decarboxylase (HemE) is important due to its location at the first branch-point in tetrapyrrole biosynthesis. We detected a complex formation between full-length polypeptides of HtpG and HemE by biochemical studies in vivo and in vitro. The interaction suppressed the enzyme activity, suggesting a regulatory role of HtpG in tetrapyrrole biosynthesis. 相似文献