The interaction of rat liver carbamoyl phosphate synthetase and ornithine transcarbamoylase with inner mitochondrial membranes

The intramitochondrial localization of the urea cycle enzymes, carbamoyl phosphate synthetase and ornithine transcarbamoylase, has been examined by both in vitro and in situ studies. The following three lines of evidence are presented to establish that significant fractions of the rat liver enzymes are loosely associated with the inner mitochondrial membrane: 1) when the mitochondrion is fractionated, the enzymes partition between the matrix and membrane fractions in the absence of detergent and partition solely to the matrix in the presence of detergent; 2) the purified enzymes associate with purified inner membrane preparations; and, 3) protein A-gold electron microscopic immunocytochemical analysis of rat liver sections reveals a nonrandom arrangement of the enzyme, with the maximal enzyme density adjacent to the inner mitochondrial membrane. These findings serve as the basis for novel potential mechanisms for regulation of the activity of the enzymes and provide additional evidence for the extensive organization of the mitochondrial matrix. The membrane interaction might also serve as the organizing factor for a carbamoyl phosphate synthetase-ornithine transcarbamoylase or other multienzyme complex.     

ATP and other purine nucleotides stimulate the inactivation of ornithine transcarbamylase by broken lysosomes

Calpain II, a high Ca2+-requiring form of Ca2+-dependent cysteine proteinase (EC 3.4.22.17), isolated from bovine lens was found to cleave actin and vimentin, two major cytoskeletal elements of the lens. Polyacrylamide gel electrophoresis revealed that actin (Mr 43 000) was broken down through intermediary products of approximate Mr 42 000 and 40 000, while vimentin (Mr 57 000) was rapidly cleaved into several fragments ranging from Mr 44 000 to 20 000. The cleavage was dependent on Ca2+ and could be blocked by calpastatin , a calpain-specific inhibitor. These findings suggest that calpain might play a role in age-related degradation of the lens cytoskeleton.     

Carbamoyl phosphate synthetase, ornithine transcarbamylase, and aspartate transcarbamylase activities in the pea ovary : changes with senescence of the unpollinated ovary or with fruit set induced by gibberellic Acid

Carbamoyl phosphate synthetase (CPS), ornithine transcarbamylase (OTC), and aspartate transcarbamylase (ATC) were assayed in extracts from unpollinated ovaries of Pisum sativum L. CPS and OTC activities were, per milligram protein, the highest reported in a plant tissue, representing an estimated 0.1% of the protein in the ovary. The OTC/CPS and ATC/CPS ratios were about 100 and 0.5, respectively, indicating that most of the carbamoyl phosphate is used for arginine synthesis. The weight, protein content, and CPS, OTC, and ATC activities per ovary were determined during the senescence of the ovary and also during fruit set induced by treatment with gibberellic acid (GA₃). In the nontreated ovary the weight and the protein first increased and then decreased dramatically, but the decrease in protein took place much earlier. In the GA₃-treated ovaries the increase in weight was considerably greater than the increase in the protein. Whether or not the ovaries were treated with GA₃, CPS, OTC, and ATC activities closely followed the changes in protein, and thus their ratios and specific activities remained essentially constant. It appears that treatment with GA₃ increases the amount of protein and enzymic activities by preventing a large increase in the rate of protein degradation. In addition, the effects of acetylglutamate, ornithine, and UMP on CPS activity were studied. The pea enzyme exhibits regulatory properties intermediate between those of Escherichia coli and the ureotelic liver enzymes.     

Crystallization,characterization, and preliminary crystallographic studies of mitochondrial carbamoyl phosphate synthetase I of Rana catesbeiana

Carbamoyl phosphate synthetase I (ammonia; E C 6.3.4.16) was purified from the liver of Rana catesbeiana (bullfrog). Crystals of the protein have been obtained at 22°C by the hanging drop vapor diffusion technique, with polyethylene glycol as precipitant. Tetragonal crystals of about 0.3 × 0.3 × 0.7 mm diffract at room temperature to at least 3.5 Å using a conventional source and are stable to X-radiation for about 12 h. Therefore, these crystals are suitablefor high resolution studies. The space group is P4₁2₁2 (or its enantiomorph P4₃2₁2), with unit cell dimensions a = b = 291.6 Å and c = 189.4 Å. Density packing considerations areconsistent with the presence of 4-6 monomers (M_r of the monomer, 160,000) in the asymmetric unit. Amino-terminal sequence of the enzyme and of a chymotryptic fragment of 73.7 kDa containing the COOH-terminus has been obtained. The extensive sequence identity with rat and human carbamoyl phosphate synthetase I indicates the relevance for mammals of structural data obtained with the frog enzyme. © 1995 Wiley-Liss, Inc.     

Mechanism of mitochondrial carbamoyl-phosphate synthetase: synthesis and properties of active CO2, precursor of carbamoyl phosphate.

This paper demonstrates the formation of "active CO2" (CO2-P), a precursor of carbamoyl phosphate (CP), with frog liver carbamoyl-phosphate synthetase. Absence of ammonia is essential for the demonstration by pulse incubation with H14CO3- of CO2-P. Adenosine triphosphate (ATP) and acetylglutamate are required for the synthesis of CO2-P, which is highly unstable in aqueous solutions (t1/2 = 0.75 s at 24 degrees C at neutral pH). In the absence of ammonia, CO2-P attains rapidly a steady-state level, which depends on the concentration of ATP and HCO3-. The "apparent KM'S" are approximately equal to those found for the adenosine triphosphate (ATPase) activity of the enzyme. The maximum level of CO2-P is limited by the amount of enzyme, and approximates 4 mol of intermediate/mol of enzyme. The unprotonated form of ammonia seems to be the species reacting with CO2-P to produce CP. The reaction of CO2-P and NH3 is very fast (rate constant kn = 8 x 10(4) M-1 S-1) and does not consume free ATP. Therefore, the 2 mol of ATP necessary for CP synthesis binds or reacts with the enzyme and/or CO2 prior to reaction with NH3. The reaction of CO2-P with NH3 also takes place in acetone under conditions at which the enzyme is not active, suggesting little or no assistance from enzyme catalysis or that a part of the catalytic site is "frozen" by the solvent in the active conformation. In the light of these and other findings, a new scheme is proposed for the mechanism of frog liver carbamoyl-phosphate synthetase and some considerations are made on the chemical nature of the intermediate and on the possible evolutionary significance of the reaction of CO2-P with NH3 in acetone.     

The analogous mechanisms of enzymatic inactivation induced by ascorbate and superoxide in the presence of copper

The mechanism of enzymatic inactivation of purified and membrane-bound acetylcholine esterase by ascorbate and copper was investigated. While the exposure of the enzyme to either ascorbate or copper did not cause enzymatic inactivation, the incubation of the enzyme with a combination of both ascorbate and copper resulted in a loss in acetylcholine esterase activity, which was time dependent. The enzymatic inactivation required either molecular oxygen or hydrogen peroxide under anaerobic conditions. Scavengers of hydroxyl radicals at concentrations of up to 100 mM did not provide protection to acetylcholine esterase. Only mannitol at very high concentrations (above 1 M) efficiently prevented the inactivation of the enzyme. The kinetics of the aerobic oxidation of reduced ascorbate in the presence of acetylcholine esterase and copper closely followed the rate of enzyme inactivation. Addition of the chelating agents EDTA and diethylenetriaminepentaacetic acid prevented both the oxidation of ascorbate and the inactivation of the enzyme. In the presence of low concentrations of histidine (0.5-2.0 mM), which forms high affinity complexes with copper, the rate of ascorbate oxidation was similar to that recorded in its absence. On the other hand, no enzyme inactivation was indicated in the presence of histidine. Low temperature EPR measurements have demonstrated the binding of copper to the enzyme, and have shown the reduction of the cupric enzyme to the corresponding cuprous complex. In view of these results, a general "site-specific" mechanism for biological damage can be offered, in which copper(II) ions are bound to enzymes or other biological macromolecules. Ascorbate plays a dual role: it reduces the cupric complex to the corresponding cuprous state and serves as a source for H2O2, which, in turn, reacts with the reduced copper complex, in a Fenton reaction. In this reaction, secondary hydroxyl radicals are site specifically formed, and react preferentially with the protein, at the site of their formation, causing its inactivation. This mechanism is analogous to that previously proposed (Samuni, A., Chevion, M., and Czapski, G. (1981) J. Biol. Chem. 256, 12632-12635) for the enhancement of the biological damage caused by superoxide in the presence of copper.     

Inhibition of Fe2+- and Fe3+- induced hydroxyl radical production by the iron-chelating drug deferiprone

Deferiprone (L1) is an effective iron-chelating drug that is widely used for the treatment of iron-overload diseases. It is known that in aqueous solutions Fe²⁺ and Fe³⁺ ions can produce hydroxyl radicals via Fenton and photo-Fenton reactions. Although previous studies with Fe²⁺ have reported ferroxidase activity by L1 followed by the formation of Fe³⁺ chelate complexes and potential inhibition of Fenton reaction, no detailed data are available on the molecular antioxidant mechanisms involved. Similarly, in vitro studies have also shown that L1–Fe³⁺ complexes exhibit intense absorption bands up to 800 nm and might be potential sources of phototoxicity. In this study we have applied an EPR spin trapping technique to answer two questions: (1) does L1 inhibit the Fenton reaction catalyzed by Fe²⁺ and Fe³⁺ ions and (2) does UV–Vis irradiation of the L1–Fe³⁺ complex result in the formation of reactive oxygen species. PBN and TMIO spin traps were used for detection of oxygen free radicals, and TEMP was used to trap singlet oxygen if it was formed via energy transfer from L1 in the triplet excited state. It was demonstrated that irradiation of Fe³⁺ aqua complexes by UV and visible light in the presence of spin traps results in the appearance of an EPR signal of the OH spin adduct (TMIO–OH, a(N)=14.15 G, a(H)=16.25 G; PBN–OH, a(N)=16.0 G, a(H)=2.7 G). The presence of L1 completely inhibited the OH radical production. The mechanism of OH spin adduct formation was confirmed by the detection of methyl radicals in the presence of dimethyl sulfoxide. No formation of singlet oxygen was detected under irradiation of L1 or its iron complexes. Furthermore, the interaction of L1 with Fe²⁺ ions completely inhibited hydroxyl radical production in the presence of hydrogen peroxide. These findings confirm an antioxidant targeting potential of L1 in diseases related to oxidative damage.     

Oxidative inactivation of carbamoyl phosphate synthetase (ammonia). Mechanism and sites of oxidation, degradation of the oxidized enzyme, and inactivation by glycerol, EDTA, and thiol protecting agents.

Acetylglutamate and ATP accelerate the oxidative inactivation of carbamoyl phosphate synthetase I by mixtures of Fe3+, ascorbate, and O2, but the mechanism of the inactivation differs with each ligand. In the presence of acetylglutamate, MgATP prevents, Mg2+, Mn2+, and catalase have no effect, and EDTA increases the inactivation, and the two phosphorylation steps of the enzyme reaction are lost simultaneously. The inactivation appears to be mediated by dehydroascorbate and is associated with the reversible oxidation of the highly reactive cysteines 1327 and 1337 and with oxidation of non-thiolic groups in the second 40-kDa domain (the enzyme consists of 4 domains of 40, 40, 60, and 20 kDa, from the amino terminus). The data are consistent with oxidation of groups at or near the site for ATPA (ATPA yields Pi; ATPB yields carbamoyl phosphate), and with the location of this site at the interphase between the second 40-kDa and the COOH-terminal domains. The oxidative inactivation promoted by ATP is inhibited by Mg2+, Mn2+, catalase, and EDTA, is not mediated by dehydroascorbate, and is not associated with oxidation of cysteines 1327 and 1337. Groups in the 60-kDa domain are oxidized. The phosphorylation step involving ATPB is lost preferentially, and the inactivation and the binding of ATPB exhibit the same dependency on the concentration of ATP. The results indicate that the oxidation is catalyzed by FeATP bound at the site for ATPB and support the binding of ATPB in the 60-kDa domain. We also demonstrate that mercaptoethanol, reducing impurities in glycerol, and dithioerythritol, in the presence of EDTA, replace ascorbate in the oxidative system. In addition, we study the influence of the oxidation on the degradation of the enzyme by rat liver lysosomes, mitochondria, and cytosol.     

Sequential up-regulation of thyroid hormone β receptor,ornithine transcarbamylase,and carbamyl phosphate synthetase mRNAs in the liver of Rana catesbeiana tadpoles during spontaneous and thyroid hormone-induced metamorphosis

During both spontaneous and thyroid hormone (TH)-induced metamorphosis, the Rana catesbeiana tadpole undergoes postembryonic developmental changes in its liver which are necessary for its transition from an ammonotelic larva to a ureotelic adult. Although this transition ultimately results from marked increases in the activities and/or de novo synthesis of the urea cycle enzymes, the precise molecular means by which TH exerts this tissue-specific response are presently unknown. Recent reports, using RNA from whole Xenopus laevis tadpole homogenates and indirect means of measuring TH receptor (TR) mRNAs, suggest a correlation between the up-regulation of TRβ-mRNAs and the general morphological changes occurring during amphibian metamorphosis. To assess whether or not this same relationship exists in a TH-responsive tissue, such as liver, we isolated and characterized a cDNA clone containing the complete nucleotide sequence for a R. catesbeiana urea cycle enzyme, ornithine transcarbamylase (OTC), as well as a genomic clone containing a portion of the hormone-binding domain of a R. catesbeiana TRβ gene. Through use of these homologous sequences and a heterologous cDNA fragment encoding rat carbamyl phosphate synthetase (CPS), we directly determined the relative levels of the TRβ, OTC, and CPS mRNAs in liver from spontaneous and TH-induced tadpoles. Our results establish that TH affects an up-regulation of mRNAs for its own receptor prior to up-regulating CPS and OTC mRNAs. Moreover, results with cultured tadpole liver demonstrate that TH, in the absence of any other hormonal influence, can affect an up-regulation of both the TRβ and OTC mRNAs. © 1992 Wiley-Liss, Inc.     

Metabolic channeling of carbamoyl phosphate, a thermolabile intermediate: evidence for physical interaction between carbamate kinase-like carbamoyl-phosphate synthetase and ornithine carbamoyltransferase from the hyperthermophile Pyrococcus furiosus

Two different approaches provided evidence for a physical interaction between the carbamate kinase-like carbamoyl-phosphate synthetase (CKase) and ornithine carbamoyltransferase (OTCase) from the hyperthermophilic archaeon Pyrococcus furiosus. Affinity electrophoresis indicated that CKase and OTCase associate into a multienzyme cluster. Further evidence for a biologically significant interaction between CKase and OTCase was obtained by co-immunoprecipitation combined with formaldehyde cross-linking experiments. These experiments support the hypothesis that CKase and OTCase form an efficient channeling cluster for carbamoyl phosphate, an extremely thermolabile and potentially toxic metabolic intermediate. Therefore, by physically interacting with each other, CKase and OTCase prevent the thermodenaturation of carbamoyl phosphate in the aqueous cytoplasmic environment.     

Interactions of glycolate-, HCO 3 - -, Cl--, and H+-balance of Scenedesmus obliquus

Excretion and absorption of glycolate by young cells of Scenedesmus obliquus (Turp.) Krüger strain D₃ grown synchronously with 2% CO₂ was compared after no pretreatment with air (CO₂-adapted) or after a 2 h adaptation to normal air (0.03% CO₂) (air-adapted). At 21% O₂, excretion occurred only from CO₂-adapted cells at high pH (pH 8.0). Under conditions where no excretion occurred, external glycolate (0.2 mM) was taken up by both air-and CO₂-adapted cells at a much faster rate at pH 5 than at pH 8. The uptake was accompanied by an apparent stoichiometric uptake of H⁺. CO₂-adapted algae exhibited high uptake rates that were even higher in the dark than in the light. Air-adapted algae showed high uptake rates in the light but only minimal uptake in the dark. The uptake rate was decreased to about 1/3 with 5% CO₂, except with CO₂-adapted cells in the light, in which a slight stimulation occurred. Cl^- ions inhibited glycolate uptake by air-adapted cells in the light; conversely, light-stimulated Cl^- uptake of these cells was inhibited by glycolate. A hypothesis is discussed according to which the internal pH regulates the uptake and release of Cl^-, HCO ₃ ^- , and glycolate.Abbreviations DCMU 3-(3,4 dichlorophenyl)-1, 1-dimethyl urea - FCCP carbonyl cyanide p-trifluoro-methoxyphenylhydrazone - HEPES 2-(4-(2-hydroxyethyl)-piperazinyl) ethanesulfonic acid - HPMS -hydroxypyridinemethanesulfonate - MES 2-morpholinoethanesulfonic acid - PCV packed cell volume     

Glycolate excretion by Anacystis nidulans: effect of HCO3- concentration, oxygen concentration and light intensity

Anacystis nidulans exhibited a light-dependent glycolate excretionwhich occurred at a maximum rate in the absence of added HCO₃^–and in the presence of high oxygen concentration. At high lightintensity excretion was linear from the beginning of the illuminationperiod. Low light intensity simply delayed the onset of excretionbut did not affect the rate of excretion once it began. Glycolateexcretion was sensitive to photosynthesis inhibitors but wasinsensitive to isonicotinyl hydrazide. (Received September 19, 1972; )     

Sequential up-regulation of thyroid hormone beta receptor, ornithine transcarbamylase, and carbamyl phosphate synthetase mRNAs in the liver of Rana catesbeiana tadpoles during spontaneous and thyroid hormone-induced metamorphosis.

During both spontaneous and thyroid hormone (TH)-induced metamorphosis, the Rana catesbeiana tadpole undergoes postembryonic developmental changes in its liver which are necessary for its transition from an ammonotelic larva to a ureotelic adult. Although this transition ultimately results from marked increases in the activities and/or de novo synthesis of the urea cycle enzymes, the precise molecular means by which TH exerts this tissue-specific response are presently unknown. Recent reports, using RNA from whole Xenopus laevis tadpole homogenates and indirect means of measuring TH receptor (TR) mRNAs, suggest a correlation between the up-regulation of TR beta-mRNAs and the general morphological changes occurring during amphibian metamorphosis. To assess whether or not this same relationship exists in a TH-responsive tissue, such as liver, we isolated and characterized a cDNA clone containing the complete nucleotide sequence for a R. catesbeiana urea cycle enzyme, ornithine transcarbamylase (OTC), as well as a genomic clone containing a portion of the hormone-binding domain of a R. catesbeiana TR beta gene. Through use of these homologous sequences and a heterologous cDNA fragment encoding rat carbamyl phosphate synthetase (CPS), we directly determined the relative levels of the TR beta, OTC, and CPS mRNAs in liver from spontaneous and TH-induced tadpoles. Our results establish that TH affects an up-regulation of mRNAs for its own receptor prior to up-regulating CPS and OTC mRNAs. Moreover, results with cultured tadpole liver demonstrate that TH, in the absence of any other hormonal influence, can affect an up-regulation of both the TR beta and OTC mRNAs.     

Effect of the combined action of flavonoids, ascorbate and alpha-tocopherol on peroxidation of phospholipid liposomes induced by Fe2+ ions

The effect of alpha-tocopherol, ascorbate, rutin and dihydroquercetin on chemiluminescence (CL) accompanying the Fe2+-induced peroxidation of unsaturated fatty acids in phospholipid liposomes has been investigated. The amplitude of CL decreased and the latent period increased in the presence of alpha-tocopherol, rutin and dihydroquercetin which is typical of peroxide radical traps. Ascorbate also reduced the CL amplitude but only at small concentrations up to about 4 microM. A further increase of ascorbate concentration had a negligible effect on the amplitude. At the same time, the latent period in CL development increased with the growth of ascorbate concentration, apparently, as a result of recycling of divalent iron oxidized in the course of lipid peroxidation. The effects of rutin and dihydroquercetin on the liposomal CL in the presence of alpha-tocopherol and ascorbate in all experiments were almost the same as when these compounds were added individually. The antioxidant effects were merely summed up without any mutual enhancement or inhibition of each other's action.     

Inactivation of Ca2(+)-, Na+K(+)-, and H+K(+)-ATPases with a carbodiimide derivative of ATP

The gamma-P adduct of ATP with 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (ATP-EDC) was synthesized and incubated with the Ca-ATPase of sarcoplasmic reticulum with the result that time-dependent complete loss of the enzyme's activity occurred. The inactivation required calcium and magnesium while ATP had a protective effect. ATP-EDC incubation with the NaK-ATPase and HK-ATPase produced partial (greater than 50%) inactivation, but had no effect on myosin S1, pyruvate kinase and hexokinase, suggesting that this ATP analog is a specific inactivator of the so-called 'P-type' ATPases.     

活性氧、NO和线粒体ATP敏感钾通道在TNF-α预处理对缺血/再灌注心肌保护中的作用

目的: 观察TNF-α预处理对缺血/再灌注心脏功能和酶学指标的影响及其可能机制.方法: 采用心脏Langendorff灌流模型.结果:与单独缺血/再灌注组相比,TNF-α(104U/L)预处理明显减弱缺血/再灌注对左室发展压、左室舒张末压、最大收缩/舒张速率和左室发展压与心率乘积的抑制作用(P<0.05),并显著降低复灌后冠脉流出液中乳酸脱氢酶(LDH)含量,增加线粒体中锰超氧化物歧化酶(Mn-SOD)活性(P<0.05);分别使用抗氧化剂2-MPG(0.3 mmol/L)、一氧化氮合酶抑制剂L-NAME(0.5 mmol/L)或线粒体ATP敏感钾通道抑制剂5-HD(100 μmol/L)预处理,减弱了TNF-α改善缺血/再灌注后心功能、抑制心肌LDH释放和诱导Mn-SOD活性增高的作用.结论: TNF-α预处理具有减轻心脏缺血/再灌注损伤的作用,这一作用可能与其诱导Mn-SOD活性增高有关,活性氧、一氧化氮和线粒体ATP敏感钾通道参与介导TNF-α的心肌保护作用.     

Estradiol protects against ATP depletion, mitochondrial membrane potential decline and the generation of reactive oxygen species induced by 3-nitroproprionic acid in SK-N-SH human neuroblastoma cells

Mitochondria are recognized as modulators of neuronal viability during ischemia, hypoxia and toxic chemical exposure, wherein mitochondria dysfunction leading to ATP depletion may be a common pathway of cell death. Estrogens have been reported to be neuroprotective and proposed to play a role in the modulation of cerebral energy/glucose metabolism. To address the involvement of 17beta-estradiol preservation of mitochondrial function, we examined various markers of mitochondrial activity in human SK-N-SH neuroblastoma cells exposed to 3-nitroproprionic acid (3-NPA), a succinate dehydrogenase inhibitor which uncouples oxidative phosphorylation. 3-NPA (10 mM) significantly increased ATP levels at 2 h then caused a 40% and a 50% decrease in ATP levels from baseline when treated for 12 h and 24 h, respectively. 3-NPA also induced significant increases in levels of cellular hydrogen peroxide and peroxynitrite at 2 h and a 60% decrease in mitochondrial membrane potential (MMP) at 12 h exposure. 17beta-Estradiol (17beta-E(2)) pretreatment restored the ATP level back to 80% at 12 h of that in control cells treated with 3-NPA but without E(2), blunted the effect of 3-NPA on MMP and reactive oxygen species levels. The present study indicates that 17beta-E(2) can preserve mitochondrial function in the face of inhibition of oxidative phosphorylation.     

Oxidative damage to bovine serum albumin induced by hydroxyl radical generating systems of xanthine oxidase + EDTA-Fe3+ and ascorbate + EDTA-Fe3+.

Oxidative damage to bovine serum albumin (BSA) was induced by hydroxyl radical (HO.) generating systems of xanthine oxidase (XO) + EDTA-Fe3+ and ascorbate + EDTA-Fe3+. Formation of bityrosine and loss of tryptophan were observed in the ascorbate + EDTA-Fe3+ system and carbonyl formation was induced by both systems. Mannitol and ethanol very strongly inhibited the carbonyl and/or bityrosine formation, indicating that the oxidative damage to BSA was due to HO(.). The sulfhydryl (SH) groups of BSA were very sensitive to the XO + EDTA-Fe3+ but not to the ascorbate + EDTA-Fe3+ system. Catalase but not hydroxyl radical scavengers or superoxide dismutase strongly inhibited the loss of SH groups, indicating that H2O2 is involved in their oxidation. Fragmentation of BSA was observed during exposure to the XO + EDTA-Fe3+ and ascorbate + EDTA-Fe3+ systems and the products presented a broad band on sodium dodecyl sulfate polyacrylamide gel electrophoresis. Little formation of amine groups was observed in these systems, indicating that little peptide bond cleavage occurred. BSA exposed to the ascorbate + EDTA-Fe3+ system was more readily degraded by trypsin than that exposed to the XO + EDTA-Fe3+ system. Elastase degraded BSA exposed to the ascorbate + EDTA-Fe3+ system but not to the XO + EDTA-Fe3+ system.     

不同来源小球藻对岩溶水Ca2+、HCO3-利用的初步研究

以外源小球藻和岩溶区筛选出的土著小球藻为研究对象, 在封闭体系中比较研究了两种不同来源小球藻对典型岩溶水中Ca2+、HCO3-的利用、藻细胞数量与其对Ca2+、HCO3-的利用率的关系和体系pH的变化。结果表明, 土著小球藻利用Ca2+、HCO3-的能力强于外源小球藻, 但外源小球藻对Ca2+的利用量高于土著小球藻, 而二者对HCO3-的利用量相同, 并且外源小球藻能够以胞外CaCO3形式产生沉淀, 而土著藻则不能形成沉淀。其次两体系中pH的变化显示, 两种小球藻光合作用都是先以水体中CO2为光合作用碳源, 然后利用HCO3-。外源小球藻能将岩溶水中29.648%的HCO3-吸收, 而土著藻能将40.625%的HCO3-通过其在食物链中的初级生产地位将岩溶碳汇转化进入到生态系统, 表现为净碳汇效应。       

Response of lupins (Lupinus angustifolius L.) and peas (Pisum sativum L.) to Fe deficiency induced by low concentrations of Fe in solution or by addition of HCO3 -

Lupins appear to be more sensitive than peas to Fe deficiency. However, when grown in nutrient solutions between pH 5–6, little difference existed between them in their ability to acidify the solution or to release Fe^III reducing compounds. This experiment was aimed at determining whether differences between species which occurred when Fe deficiency was induced by withholding Fe from an acid solution, are maintained when Fe deficiency is induced by addition of HCO₃ ^-. Lupins and peas were grown in nutrient solutions at 0, 2 and 6 μM of Fe^III EDDHA and either with or without HCO₃ ^- (6 mM). Bicarbonate induced symptoms of Fe deficiency (chlorosis) in both lupins and peas, and markedly decreased the growth of shoots. Symptoms appeared sooner and were more severe in lupins than in peas. Growing plants without HCO₃ ^-, but at the lowest Fe level, decreased the growth and Fe concentration of shoots of lupins but did not induce chlorosis. Growing peas in this treatment, decreased Fe concentrations, but to a lesser extent than in lupins, and did not decrease growth. H⁺-ion extrusion and release of Fe^III reducing compounds was greater in lupins than in peas. Bicarbonate also decreased the growth of roots of lupins but increased the growth of roots of peas. Results indicate that when Fe deficiency is induced by HCO₃ ^-, then the response of lupins and peas are similar to their response in acid solution culture. Differences between species therefore could not be explained by their relative abilities to acidify or release Fe^III reducing compounds. Greater control of the distribution of Fe within the shoots, the presence of a pool of Fe within the roots, a lower threshold for Fe uptake, or a higher content of seed-Fe, may therefore be the reason for the lower sensitivity of peas than lupins to Fe deficiency.