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41.
1. The enzyme–substrate complex of yeast cytochrome c peroxidase is used as a sensitive, specific and accurate spectrophotometric H2O2 indicator. 2. The cytochrome c peroxidase assay is suitable for use with subcellular fractions from tissue homogenates as well as with pure enzyme systems to measure H2O2 generation. 3. Mitochondrial substrates entering the respiratory chain on the substrate side of the antimycin A-sensitive site support the mitochondrial generation of H2O2. Succinate, the most effective substrate, yields H2O2 at a rate of 0.5nmol/min per mg of protein in state 4. H2O2 generation is decreased in the state 4→state 3 transition. 4. In the combined mitochondrial–peroxisomal fraction of rat liver the changes in the mitochondrial generation of H2O2 modulated by substrate, ADP and antimycin A are followed by parallel changes in the saturation of the intraperoxisomal catalase intermediate. 5. Peroxisomes supplemented with uric acid generate extraperoxisomal H2O2 at a rate (8.6–16.4nmol/min per mg of protein) that corresponds to 42–61% of the rate of uric acid oxidation. Addition of azide increases these H2O2 rates by a factor of 1.4–1.7. 6. The concentration of cytosolic uric acid is shown to vary during the isolation of the cellular fractions. 7. Microsomal fractions produce H2O2 (up to 1.7nmol/min per mg of protein) at a ratio of 0.71–0.86mol of H2O2/mol of NADP+ during the oxidation of NADPH. H2O2 is also generated (6–25%) during the microsomal oxidation of NADH (0.06–0.025mol of H2O2/mol of NAD+). 8. Estimation of the rates of production of H2O2 under physiological conditions can be made on the basis of the rates with the isolated fractions. The tentative value of 90nmol of H2O2/min per g of liver at 22°C serves as a crude approximation to evaluate the biochemical impact of H2O2 on cellular metabolism. 相似文献
42.
Two plant-type ferredoxins were isolated and purified from a blue-green alga, Nostoc verrucosum. They were separable by chromatography on a DEAE-cellulose column. The slow-moving band was designated ferredoxin I (Fd I) and the fast-moving band was ferredoxin II (Fd II). The ratio of the yield of ferredoxins I and II was about 1 : 0.84. Both ferredoxins had absorption spectra similar to those of plant-type ferredoxins. Two atoms of non-heme iron and two of labile sulfur were found per mol of both ferredoxin I and ferredoxin II. Their molecular weights were identical and estimated to be about 18 000 by a gel filtration method. The biochemical activities of these Nostoc ferredoxins were studied: the NADP photoreduction activity on one hand and the NADP-cytochrome c reductase activity on the other. 相似文献
43.
Łukasz Wejnerowski Ewa Poniecka Jakub Buda Piotr Klimaszyk Agnieszka Piasecka Marcin Krzysztof Dziuba Gianmarco Mugnai Nozomu Takeuchi Krzysztof Zawierucha 《Journal of phycology》2023,59(5):939-949
Cryoconite, the dark sediment on the surface of glaciers, often aggregates into oval or irregular granules serving as biogeochemical factories. They reduce a glacier's albedo, act as biodiversity hotspots by supporting aerobic and anaerobic microbial communities, constitute one of the organic matter (OM) sources on glaciers, and are a feeder for micrometazoans. Although cryoconite granules have multiple roles on glaciers, their formation is poorly understood. Cyanobacteria are ubiquitous and abundant engineers of cryoconite hole ecosystems. This study tested whether cyanobacteria may be responsible for cryoconite granulation as a sole biotic element. Incubation of Greenlandic, Svalbard, and Scandinavian cyanobacteria in different nutrient availabilities and substrata for growth (distilled water alone and water with quartz powder, furnaced cryoconite without OM, or powdered rocks from glacial catchment) revealed that cyanobacteria bind mineral particles into granules. The structures formed in the experiment resembled those commonly observed in natural cryoconite holes: they contained numerous cyanobacterial filaments protruding from aggregated mineral particles. Moreover, all examined strains were confirmed to produce extracellular polymeric substances (EPS), which suggests that cryoconite granulation is most likely due to EPS secretion by gliding cyanobacteria. In the presence of water as the only substrate for growth, cyanobacteria formed mostly carpet-like mats. Our data empirically prove that EPS-producing oscillatorialean cyanobacteria isolated from the diverse community of cryoconite microorganisms can form granules from mineral substrate and that the presence of the mineral substrate increases the probability of the formation of these important and complex biogeochemical microstructures on glaciers. 相似文献
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We investigated the effects of metabolic inhibition on both the shift in the equilibrium potential for Cl(-) (E(Cl)) and the run down of GABA(A) receptor responses, using nystatin- and gramicidin-perforated patch-clamp recordings from rat hippocampal CA1 neurons. Metabolic inhibition with NaCN decreased outward GABAergic currents while increasing inward GABAergic currents. E(Cl) showed a positive shift almost immediately after metabolic poisoning. This shift always occurred prior to GABA receptor run down, which was observed as decreases in whole cell conductance during application of a GABA(A) receptor agonist. The results indicate that GABAergic responses tend to become depolarizing during metabolic inhibition and the run down of the GABAergic response may therefore be neuroprotective against excitotoxicity. Furthermore the results illustrate the importance of considering both changes in receptor function and current driving force, and their temporal relationship, in order to understand the physiological response of the GABAergic system during metabolic stress. 相似文献
46.
Nagae M Nishi N Murata T Usui T Nakamura T Wakatsuki S Kato R 《The Journal of biological chemistry》2006,281(47):35884-35893
The galectins are a family of beta-galactoside-binding animal lectins with a conserved carbohydrate recognition domain (CRD). They have a high affinity for small beta-galactosides, but binding specificity for complex glycoconjugates varies considerably within the family. The ligand recognition is essential for their proper function, and the structures of several galectins have suggested their mechanism of carbohydrate binding. Galectin-9 has two tandem CRDs with a short linker, and we report the crystal structures of mouse galectin-9 N-terminal CRD (NCRD) in the absence and the presence of four ligand complexes. All structures form the same dimer, which is quite different from the canonical 2-fold symmetric dimer seen for galectin-1 and -2. The beta-galactoside recognition mechanism in the galectin-9 NCRD is highly conserved among other galectins. In the apo form structure, water molecules mimic the ligand hydrogen-bond network. The galectin-9 NCRD can bind both N-acetyllactosamine (Galbeta1-4GlcNAc) and T-antigen (Galbeta1-3GalNAc) with the proper location of Arg-64. Moreover, the structure of the N-acetyllactosamine dimer (Galbeta1-4GlcNAcbeta1-3Galbeta1-4GlcNAc) complex shows a unique binding mode of galectin-9. Finally, surface plasmon resonance assay showed that the galectin-9 NCRD forms a homophilic dimer not only in the crystal but also in solution. 相似文献
47.
Kamei N Tobe K Suzuki R Ohsugi M Watanabe T Kubota N Ohtsuka-Kowatari N Kumagai K Sakamoto K Kobayashi M Yamauchi T Ueki K Oishi Y Nishimura S Manabe I Hashimoto H Ohnishi Y Ogata H Tokuyama K Tsunoda M Ide T Murakami K Nagai R Kadowaki T 《The Journal of biological chemistry》2006,281(36):26602-26614
Adipose tissue expression and circulating concentrations of monocyte chemoattractant protein-1 (MCP-1) correlate positively with adiposity. To ascertain the roles of MCP-1 overexpression in adipose, we generated transgenic mice by utilizing the adipocyte P2 (aP2) promoter (aP2-MCP-1 mice). These mice had higher plasma MCP-1 concentrations and increased macrophage accumulation in adipose tissues, as confirmed by immunochemical, flow cytometric, and gene expression analyses. Tumor necrosis factor-alpha and interleukin-6 mRNA levels in white adipose tissue and plasma non-esterified fatty acid levels were increased in transgenic mice. aP2-MCP-1 mice showed insulin resistance, suggesting that inflammatory changes in adipose tissues may be involved in the development of insulin resistance. Insulin resistance in aP2-MCP-1 mice was confirmed by hyperinsulinemic euglycemic clamp studies showing that transgenic mice had lower rates of glucose disappearance and higher endogenous glucose production than wild-type mice. Consistent with this, insulin-induced phosphorylations of Akt were significantly decreased in both skeletal muscles and livers of aP2-MCP-1 mice. MCP-1 pretreatment of isolated skeletal muscle blunted insulin-stimulated glucose uptake, which was partially restored by treatment with the MEK inhibitor U0126, suggesting that circulating MCP-1 may contribute to insulin resistance in aP2-MCP-1 mice. We concluded that both paracrine and endocrine effects of MCP-1 may contribute to the development of insulin resistance in aP2-MCP-1 mice. 相似文献
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49.
Mamoru Tamura Nozomu Oshino Britton Chance Ian A. Silver 《Archives of biochemistry and biophysics》1978,191(1):8-22
The optical characteristics of hemoglobin-free perfused rat heart have been examined in detail. Ethyl hydrogen peroxide is found to convert myoglobin into “ferryl compound” in the perfused heart, as is also seen in vitro. After pretreatment with ethyl hydrogen peroxide, a typical mitochondrial absorption spectrum, similar to that of isolated rat heart mitochondria, is obtained in perfused heart. The overall absorption spectrum of the heart obtained by the aerobic to anaerobic transition is a superposition of the mitochondrial spectrum on that of myoglobin. By comparing these spectra, it is found that measurement of cytochrome a + a3 at 605–620 nm is possible in spite of the absorbance change due to the oxygenation-deoxygenation of myoglobin, whereas the wavelength pairs for cytochrome c at 550-540 nm, cytochrome b at 562–575 nm and cytochrome a + a3 at 445–450 nm can not be used in the heart because of interference from the absorption change of myoglobin. The partial pressure of O2 (P50) which is required for half maximal deoxygenation (or oxygenation) of myoglobin in perfused heart is found to be 2.4 mm Hg at room temperature and the Hill constant, n, is 1.1; these values are similar to those of myoglobin purified from rat heart. The steady-state O2 titration has been performed by using absorbancy changes of myoglobin and cytochrome a + a3 as intracellular O2 indicators. In the perfused heart, the percentage change of oxygenation-deoxygenation of myoglobin parallels the oxidation-reduction of cytochrome a + a3, while the mixture of purified myoglobin and isolated mitochondria shows a deviation, reflecting the difference of O2 affinities between myoglobin and cytochrome a + a3. The results indicate that there may be an O2 gradient between cytosolic and mitochondrial compartments in the hemoglobin-free perfused heart. The absorption changes of myoglobin and of cytochrome a + a3 can be measured in a single contraction-relaxation cycle. A triple beam method was introduced to eliminate the effect of light scattering changes in these measurements. The results demonstrated that myoglobin is more oxygenated during the systolic and diastolic periods and deoxygenated in the resting period, whereas cytochrome a + a3 is more reduced in systole and diastole and oxidized in the resting state. Changing the perfusion conditions greatly alters the time course of the events which occur during the contraction-relaxation cycle of the perfused heart. 相似文献
50.