74 research outputs found
Implications of adenylate kinase-governed equilibrium of adenylates on contents of free magnesium in plant cells and compartments
Involvement of cyanide-resistant and rotenone-insensitive pathways of mitochondrial electron transport during oxidation of glycine in higher plants
AbstractMetabolism of glycine in isolated mitochondria and protoplasts was investigated in photosynthetic, etiolated (barley and pea leaves) and fat-storing (maize scutellum) tissues using methods of [1-14C]glycine incorporation and counting of 14CO2 evolved, oxymetric measurement of glycine oxidation and rapid fractionation of protoplasts incubated in photorespiratory conditions with consequent determination of ATP/ADP ratios in different cell compartments. The involvement of different paths of electron transport in mitochondria during operation of glycine decarboxylase complex (GDC) was tested in different conditions, using aminoacetonitrile (AAN), the inhibitor of glycine oxidation in mitochondria, rotenone, the inhibitor of Complex I of mitochondrial electron transport, and inhibitors of cytochrome oxidase and alternative oxidase. It was shown that glycine has a preference to other substrates oxidized in mitochondria only in photosynthetic tissue where succinate and malate even stimulated its oxidation. Rotenone had no or small effect on glycine oxidation, whereas the role of cyanide-resistant path increased in the presence of ATP. Glycine oxidation increased ATP/ADP ratio in cytosol of barley protoplasts incubated in the presence of CO2, but not in the CO2-free medium indicating that in conditions of high photorespiratory flux oxidation of NADH formed in the GDC reaction passes via the non-coupled paths. Activity of GDC in fat-storing tissue correlated with the activity of glyoxylate-cycle enzymes, glycine oxidation did not reveal preference to other substrates and the involvement of paths non-connected with proton translocation was not pronounced. It is suggested that the preference of glycine to other substrates oxidized in mitochondria is achieved in photosynthetic tissue by switching to rotenone-insensitive intramitochrondrial NADH oxidation and by increasing of alternative oxidase involvement in the presence of glycine
Information and symmetry: Adumbrating the abstract core of complex systems
Information and symmetry are essential theoretical concepts that underlie the scientific explanation of a variety of complex systems. In spite of clear-cut developments around both concepts, their intersection is really problematic, either in fields related to mathematics, physics, and chemistry, or even more in those pertaining to biology, neurosciences, and social sciences. The present Special Issue explores recent developments, both theoretical and applied, in most of these disciplines
Plant hemoglobins: Important players at the crossroads between oxygen and nitric oxide
AbstractPlant hemoglobins constitute a diverse group of hemeproteins and evolutionarily belong to three different classes. Class 1 hemoglobins possess an extremely high affinity to oxygen and their main function consists in scavenging of nitric oxide (NO) at very low oxygen levels. Class 2 hemoglobins have a lower oxygen affinity and they facilitate oxygen supply to developing tissues. Symbiotic hemoglobins in nodules have mostly evolved from class 2 hemoglobins. Class 3 hemoglobins are truncated and represent a clade with a very low similarity to class 1 and 2 hemoglobins. They may regulate oxygen delivery at high O2 concentrations. Depending on their physical properties, hemoglobins belong either to hexacoordinate non-symbiotic or pentacoordinate symbiotic groups. Plant hemoglobins are plausible targets for improving resistance to multiple stresses
NO-degradation by alfalfa class 1 hemoglobin (Mhb1): a possible link to PR-1a gene expression in Mhb1-overproducing tobacco plants
AbstractTobacco plants overproducing alfalfa class 1 hemoglobin (HOT plants) have been shown to have reduced necrotic symptom development. Here, we show that this altered pathogenic response is linked to a significant increase in the nitric oxide (NO)-affected pathogenesis-related (PR-1a) transcript accumulation in the transgenic plants. Homogenates of HOT transgenic seedlings were also found to have higher NO-scavenging activity than non-transformed ones. The NO-scavenging properties of recombinant alfalfa class1 hemoglobin have been examined. Recombinant Mhb1 (rMhb1) was produced in bacteria and purified using polyethylene glycol (10–25%) fractionation, chromatography on DEAE–Sephacel, and Phenyl Superose columns. After the final purification step, the obtained preparations were near homogeneous and had a molecular weight of 44 kDa determined by size-exclusion chromatography and 23 kDa by SDS–PAGE, indicating that rMhb1 is a dimer. The protein participated in NO-degradation activity with NAD(P)H as a cofactor. After ion-exchange columns, addition of FAD was necessary for exhibiting maximal NO-degradation activity. The NAD(P)H-dependent NO-scavenging activity of rMhb1, which is similar to that of barley hemoglobin, supports a conclusion that both monocot and dicot class 1 hemoglobins can affect cellular NO levels by scavenging NO formed during hypoxia, pathogen attack and other stresses
Dihydrolipoamide dehydrogenase from porcine heart catalyzes NADH-dependent scavenging of nitric oxide
Abstract Dihydrolipoamide dehydrogenase (DLDH; EC 1.8.1.4) from porcine heart is capable of using nitric oxide (NO) as an electron acceptor, with NADH as the electron donor, forming nitrate in the reaction. NADPH was not effective as an electron donor. The reaction had a pH optimum near 6 and was not inhibited by cyanide or diphenyleneiodonium ions. The K m for NADH was 10 lM, while that for NO was 0.5 lM. The rate of NO conversion was comparable to the rate of lipoamide conversion (200 lmol min À1 mg À1 protein at pH 6). Cytochrome c or myoglobin were poor electron acceptors by themselves but, in the presence of methylene blue, DLDH had an activity of 5-7 lmol min À1 mg À1 protein with these substrates, indicating that DLDH can act also as a methemoglobin reductase. While the K m of DLDH for NO is relatively low, it is in the physiological range of NO levels encountered in the tissue. The enzyme may, therefore, have a significant role in modifying NO levels under specific cell conditions
Current approaches to measure nitric oxide in plants
Nitric oxide (NO) is now established as an important signalling molecule in plants where it influences growth, development, and responses to stress. Despite extensive research, the most appropriate methods to measure and localize these signalling radicals are debated and still need investigation. Many confounding factors such as the presence of other reactive intermediates, scavenging enzymes, and compartmentation influence how accurately each can be measured. Further, these signalling radicals have short half-lives ranging from seconds to minutes based on the cellular redox condition. Hence, it is necessary to use sensitive and specific methods in order to understand the contribution of each signalling molecule to various biological processes. In this review, we summarize the current knowledge on NO measurement in plant samples, via various methods. We also discuss advantages, limitations, and wider applications of each method
Roles for Plant Mitochondrial Alternative Oxidase Under Normoxia, Hypoxia, and Reoxygenation Conditions
Alternative oxidase (AOX) is a non-energy conserving terminal oxidase in the plant mitochondrial electron transport chain (ETC) that has a lower affinity for oxygen than does cytochrome (cyt) oxidase. To investigate the role(s) of AOX under different oxygen conditions, wild-type (WT) Nicotiana tabacum plants were compared with AOX knockdown and overexpression plants under normoxia, hypoxia (near-anoxia), and during a reoxygenation period following hypoxia. Paradoxically, under all the conditions tested, the AOX amount across plant lines correlated positively with leaf energy status (ATP/ADP ratio). Under normoxia, AOX was important to maintain respiratory carbon flow, to prevent the mitochondrial generation of superoxide and nitric oxide (NO), to control lipid peroxidation and protein S-nitrosylation, and possibly to reduce the inhibition of cyt oxidase by NO. Under hypoxia, AOX was again important in preventing superoxide generation and lipid peroxidation, but now contributed positively to NO amount. This may indicate an ability of AOX to generate NO under hypoxia, similar to the nitrite reductase activity of cyt oxidase under hypoxia. Alternatively, it may indicate that AOX activity simply reduces the amount of superoxide scavenging of NO, by reducing the availability of superoxide. The amount of inactivation of mitochondrial aconitase during hypoxia was also dependent upon AOX amount, perhaps through its effects on NO amount, and this influenced carbon flow under hypoxia. Finally, AOX was particularly important in preventing nitro-oxidative stress during the reoxygenation period, thereby contributing positively to the recovery of energy status following hypoxia. Overall, the results suggest that AOX plays a beneficial role in low oxygen metabolism, despite its lower affinity for oxygen than cytochrome oxidase
Control of Rubisco function via homeostatic equilibration of CO2 supply
Rubisco is the most abundant protein on Earth that serves as the primary engine of carbon assimilation. It is characterized by a slow rate and low specificity for CO2 leading to photorespiration. We analyze here the challenges of operation of this enzyme as the main carbon fixation engine. The high concentration of Rubisco exceeds that of its substrate CO2 by 2–3 orders of magnitude; however, the total pool of available carbon in chloroplast, i.e., mainly bicarbonate, is comparable to the concentration of Rubisco active sites. This makes the reactant stationary assumption (RSA), which is essential as a condition of satisfying the Michaelis–Menten (MM) kinetics, valid if we assume that the delivery of CO2 from this pool is not limiting. The RSA is supported by active carbonic anhydrases (CA) that quickly equilibrate bicarbonate and CO2 pools and supply CO2 to Rubisco. While the operation of stromal CA is independent of light reactions, the thylakoidal CA associated with PSII and pumping CO2 from the thylakoid lumen is coordinated with the rate of electron transport, water splitting and proton gradient across the thylakoid membrane. At high CO2 concentrations, CA becomes less efficient (the equilibrium becomes unfavorable), so a deviation from the MM kinetics is observed, consistent with Rubisco reaching its Vmax at approximately 50% lower level than expected from the classical MM curve. Previously, this deviation was controversially explained by the limitation of RuBP regeneration. At low ambient CO2 and correspondingly limited capacity of the bicarbonate pool, its depletion at Rubisco sites is relieved in that the enzyme utilizes O2 instead of CO2, i.e., by photorespiration. In this process, CO2 is supplied back to Rubisco, and the chloroplastic redox state and energy level are maintained. It is concluded that the optimal performance of photosynthesis is achieved via the provision of continuous CO2 supply to Rubisco by carbonic anhydrases and photorespiration
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