| 11. |
- Hanson, D T, et al.
(författare)
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The Chlamydomonas reinhardtii cia3 mutant lacking a thylakoid lumen-localized carbonic anhydrase is limited by CO2 supply to rubisco and not photosystem II function in vivo
- 2003
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Ingår i: Plant Physiology. - : Oxford University Press (OUP). - 0032-0889 .- 1532-2548. ; 132:4, s. 2267-2275
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Tidskriftsartikel (refereegranskat)abstract
- The Chlamydomonas reinhardtii cia3 mutant has a phenotype indicating that it requires high-CO2 levels for effective photosynthesis and growth. It was initially proposed that this mutant was defective in a carbonic anhydrase (CA) that was a key component of the photosynthetic CO2-concentrating mechanism (CCM). However, more recent identification of the genetic lesion as a defect in a lumenal CA associated with photosystem II (PSII) has raised questions about the role of this CA in either the CCM or PSII function. To resolve the role of this lumenal CA, we re-examined the physiology of the cia3 mutant. We confirmed and extended previous gas exchange analyses by using membrane-inlet mass spectrometry to monitor O-16(2), O-18(2), and CO2 fluxes in vivo. The results demonstrate that PSII electron transport is not limited in the cia3 mutant at low inorganic carbon (Ci). We also measured metabolite pools sizes and showed that the RuBP pool does not fall to abnormally low levels at low Ci as might be expected by a photosynthetic electron transport or ATP generation limitation. Overall, the results demonstrate that under low Ci conditions, the mutant lacks the ability to supply Rubisco with adequate CO2 for effective CO2 fixation and is not limited directly by any aspect of PSII function. We conclude that the thylakoid CA is primarily required for the proper functioning of the CCM at low Ci by providing an ample supply of CO2 for Rubisco.
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| 12. |
- Hiltonen, Thomas, 1960-, et al.
(författare)
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Intracellular beta-carbonic anhydrase of the unicellular green alga Coccomyxa
- 1998
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Ingår i: Plant Physiology. - 0032-0889 .- 1532-2548. ; 117:4, s. 1341-1349
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Tidskriftsartikel (refereegranskat)abstract
- Carbonic anhydrase (CA) (EC 4.2.1.1) enzymes catalyze the reversible hydration of CO,, a reaction that is important in many physiological processes. We have cloned and sequenced a full length cDNA encoding an intracellular P-CA from the unicellular green alga Coccomyxa. Nucleotide sequence data show that the isolated cDNA contains an open reading frame encoding a polypeptide of 227 amino acids. The predicted polypeptide is similar to beta-type CAs from Escherichia coli and higher plants, with an identity of 26% to 30%. The Coccomyxa cDNA was overexpressed in E. coli, and the enzyme was purified and biochemically characterized. The mature protein is a homotetramer with an estimated molecular mass of 100 kD. The CO2-hydration activity of the Coccomyxa enzyme is comparable with that of the pea homolog. However, the activity of Coccomyxa CA is largely insensitive to oxidative conditions, in contrast to similar enzymes from most higher plants. Fractionation studies further showed that Coccomyxa CA is extrachloroplastic.
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| 13. |
- Hiltonen, Thomas, 1960-, et al.
(författare)
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PURIFICATION AND CHARACTERIZATION OF AN INTRACELLULAR CARBONIC-ANHYDRASE FROM THE UNICELLULAR GREEN-ALGA COCCOMYXA
- 1995
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Ingår i: Planta. - 0032-0935 .- 1432-2048. ; 195:3, s. 345-351
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Tidskriftsartikel (refereegranskat)abstract
- An intracellular carbonic anhydrase (CA; EC 4.2.1.1) was purified and characterised from the unicellular green alga Coccomyxa sp. Initial studies showed that cultured Coccomyxa cells contain an intracellular CA activity around 100 times higher than that measured in high-CO2-grown cells of Chlamydomonas reinhardtii CW 92. Purification of a protein extract containing the CA activity was carried out using ammonium-sulphate precipitation followed by anion-exchange chromatography. Proteins were then separated by native (non-dissociating) polyacrylamide gel electrophoresis, with each individual protein band excised and assayed for CA activity. Measurements revealed CA activity associated with two discrete protein bands with similar molecular masses of 80 +/- 5 kDa. Dissociation by denaturing polyacrylamide gel electrophoresis showed that both proteins contained a single polypeptide of 26 kDa, suggesting that each 80-kDa native protein was a homogeneous trimer. Isoelectric focusing of the 80-kDa proteins also produced a single protein band at a pH of 6.5. Inhibition studies on the purified CA extract showed that 50% inhibition of CA activity was obtained using 1 mu M azetazolamide. Polyclonal antibodies against the 26-kDa CA were produced and shown to have a high specific binding to a single polypeptide in soluble protein extracts from Coccomyxa. cells. The same antiserum, however, failed to cross-react with soluble proteins isolated from two different species of green algae, Chlamydomonas reinhardtii and Chlorella vulgaris. Correspondingly, antisera directed against pea chloroplastic CA, extracellular CA from C. reinhardtii and human CAII, showed no cross-hybridisation to the 26-kDa polypeptide in Coccomyxa. The 26-kDa protein was confirmed as being a CA by N-terminal sequencing of two internal polypeptide fragments and alignment of these sequences with that of previously identified CA proteins from several different species.
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| 14. |
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| 15. |
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| 16. |
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| 17. |
- HOFSLAGARE, O, et al.
(författare)
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THE EFFECT OF ARSENATE AND ARSENITE ON PHOTOSYNTHESIS IN SCENEDESMUS-OBLIQUUS - A POTENTIOMETRIC STUDY IN A CLOSED CO2-SYSTEM
- 1994
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Ingår i: Chemical speciation and bioavailability. - 0954-2299 .- 2047-6523. ; 6:4, s. 95-102
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Tidskriftsartikel (refereegranskat)abstract
- A potentiometric titration method was used to study the adverse effect of arsenate (As(V) and arsenite (As(III) on inorganic carbon uptake in suspensions of the green alga Scenedesmus obliquus. The measurements were performed in a closed CO2-system with diluted synthetic seawater (1 parts per thousand salinity) as ionic medium. Usually, the algal chlorophyll concentration was 0.4 mg dm-3, while the arsenate- and arsenite-concentrations were varied within the limits 0.1 to 200 mumol dm-3. In some experiments arsenate toxicity was studied in the presence of 1 to 100 mumol dm-3 of phosphate (P(V)). With concentrations of arsenate or arsenite less than 0.1 mumol dm-3 no toxic effects were observed. However, at As-concentrations of 200 mumol dm-3, the algal carbon uptake was reduced by 41% with arsenate and 29% with arsenite, i.e., arsenate is more toxic to Scenedesmus obliquus than arsenite. The toxicity of arsenate was negligible in the presence of a ten fold excess of phosphate. This is probably due to chemical similarities between arsenate and phosphate causing competition between the ions for the binding sites. The importance of taking the speciation as well as the buffer capacity of the algal system into account, when calculating the carbon uptake, is also discussed.
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| 18. |
- Hägg Samuelsson, Ulrika, 1973, et al.
(författare)
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Physical exercise capacity is associated with coronary and peripheral vascular function in healthy young adults.
- 2005
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Ingår i: American journal of physiology. Heart and circulatory physiology. - : American Physiological Society. - 0363-6135 .- 1522-1539. ; 289:4, s. H1627-34
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Tidskriftsartikel (refereegranskat)abstract
- Short-term exercise training has been shown to improve cardiovascular function, whereas long-term effects of a physically active lifestyle, on coronary artery function in particular, are still not well studied. We explored possible relationships between physical exercise capacity and coronary and peripheral vascular function in healthy young adults. Twenty-nine healthy young male and female volunteers participated in the study. They underwent 1) basic clinical and echocardiographic characterization, 2) coronary flow velocity reserve (CFVR) measurement of the left anterior descending coronary artery (LAD), 3) common carotid artery (CCA) intima-media thickness (IMT) measurement, 4) assessment of CCA stiffness index (SI), 5) forearm flow-mediated vasodilation (FMD), and 6) submaximal exercise test. The calculated weight-adjusted maximal oxygen uptake capacity (Vo(2 max)(c)) was positively correlated to LAD CFVR and inversely correlated to IMT and SI. Also, subjects with high compared with moderate exercise capacity had higher FMD. In addition, subjects with LAD CFVR in the upper median had greater ratios between endothelium-dependent and -independent vasodilation in the forearm and lower SI in CCA. High exercise capacity due to a physically active lifestyle is associated with high coronary and peripheral artery function, indicating an early protective role of physical exercise for cardiovascular health.
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| 19. |
- Karlsson, Jan, 1966-, et al.
(författare)
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EFFECT OF VANADATE ON PHOTOSYNTHESIS AND THE ATP ADP RATIO IN LOW-CO2-ADAPTED CHLAMYDOMONAS-REINHARDTII CELLS
- 1994
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Ingår i: Planta. - 0032-0935 .- 1432-2048. ; 192:1, s. 46-51
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Tidskriftsartikel (refereegranskat)abstract
- We have assessed the effect of vanadate as an inhibitor of plasma-membrane ATPase on photosynthesis and the ATP/ADP ratio in Chlamydomonas reinhardtii CW-92 (a mutant strain lacking a cell wall). This effect was compared in low-CO2-adapted cells grown in media bubbled with air containing 400 or 70 muL . L-1 CO2. Evidence is presented indicating that cells grown at 70 muL . L-1 CO2 have a higher rate of photosynthetic O2 evolution than cells grown at 400 muL . L-1 CO2, at limiting carbon concentrations. Extracellular and intracellular carbonic-anhydrase activities were, however, similar in cells grown in both of the low-carbon conditions. Vanadate inhibited, to a different extent, the HCO3--dependent O2 evolution in cells grown at 400 and 70 muL . L-1 CO2. At 400 muM vanadate, inhibition reached 70-75 % in cells grown at 400 muL . L-1 but only 50 % in those grown at 70 muL . L-1 CO2. The ATP/ADP ratios determined with and without vanadate at limiting concentrations of dissolved inorganic carbon indicated that more ATP was hydrolysed in algae grown at 70 muL . L-1 than in those grown at 400 muL . L-1 CO2. We conclude that the maximal capacity to accumulate dissolved inorganic carbon is inversely related to the CO2 concentration in the medium. Activation and - or synthesis of vanadate-sensitive ATPase may be the major explanation for the higher capacity for HCO3--dependent O2 evolution in cells grown under limited CO2 concentrations.
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| 20. |
- Karlsson, Jan, 1966-, et al.
(författare)
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Intracellular carbonic anhydrase of Chlamydomonas reinhardtii
- 1995
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Ingår i: Plant Physiology. - : Oxford University Press (OUP). - 0032-0889 .- 1532-2548. ; 109:2, s. 533-539
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Tidskriftsartikel (refereegranskat)abstract
- An intracellular carbonic anhydrase (CA; EC 4.2.1.1) was purified to homogeneity from a mutant strain of Chlamydomonas reinhardtii (CW 92) lacking a cell wall. Intact cells were washed to remove periplasmic CA and were lysed and fractionated into soluble and membrane fractions by sedimentation. All of the CA activity sedimented with the membrane fraction and was dissociated by treatment with a buffer containing 200 mM KCI. Solubilized proteins were fractionated by ammonium sulfate precipitation, anionic exchange chromatography, and hydrophobic interaction chromatography. The resulting fraction had a specific activity of 1260 Wilbur-Anderson units/mg protein and was inhibited by acetazolamide (50% inhibition concentration, 12 nM). Final purification was accomplished by the specific absorption of the enzyme to a Centricon-10 microconcentrator filter. A single, 29.5-kD polypeptide was eluted from the filter with sodium dodecyl sulfate-polyacrylamide gel electrophoresis sample buffer, and a 1.5 M ammonium sulfate eluate contained CA activity. In comparison with human CA isoenzyme II, the N-terminal and internal amino acid sequences from the 29.5-kD polypeptide were 40% identical with the N-terminal region and 67% identical with an internal conserved region. Based on this evidence, we postulate that the 29.5-kD polypeptide is an internal CA in C. reinhardtii and that the enzyme is closely related to the alpha-type CAs observed in animal species.
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