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41.
Delocalized chemiosmotic coupling of oxidative phosphorylation requires that a single-value correlation exists between the extent of and the kinetic parameters of respiration and ATP synthesis. This expectation was tested experimentally in nigericin-treated plant mitochondria in single combined experiments, in which simultaneously respiration (in State 3 and in State 4) was measured polarographically, FΔψ (which under these conditions was equivalent to ) was evaluated potentiometrically from the uptake of tetraphenylphosphonium+ and the rate of phosphorylation was estimated from the transient depolarization of mitochondria during State 4-State 3-State 4 transitions. The steady-state rates of the different biochemical reactions were progressively inhibited by specific inhibitors active with different modalities on various steps of the energy-transducing process: succinate respiration was inhibited competitively with malonate or noncompetitively with antimycin A, or by limiting the rate of transport into the mitochondria of the respiratory substrate with phenylsuccinate; was dissipated by uncoupling with increasing concentrations of valinomycin; ADP phosphorylation was limited with oligomycin. The results indicate generally that when the rate of respiratory electron flow is decreased, a parallel inhibition of the rate of phosphorylation is also observed, while very limited effects can be detected on the extent of . This behavior is in marked contrast to the effect of uncoupling where the decreased rate of ATP synthesis is clearly due to energy limitation. Extending previous observations in bacterial photosynthesis and in respiration by animal mitochondria and submitochondrial particles the results indicate, therefore, that respiration tightly controls the rate of ATP synthesis, with a mechanism largely independent of . These data cannot be reconciled with a delocalized chemiosmotic coupling model. 相似文献
42.
A high recovery method for concentrating microgram quantities of protein from large volumes of solution 总被引:4,自引:0,他引:4
A method is presented for the recovery of 40-80% of the protein from a 1 microgram/ml solution. The final protein pellet is free of detergent and other ionic compounds and is thus compatible with any denaturing solution. The primary structure of the protein is unaffected by the procedure, making the final pellet an ideal sample for any analytical procedure to determine protein structure. 相似文献
43.
The preparative aspects of isoelectric focusing (IEF) in immobilized pH gradients (IPG) have been investigated as a function of the following parameters: environmental ionic strength (I), gel geometry and shape of pH gradient. As model proteins, hemoglobin (Hb) A and a minor, glycosylated component (HbA1c), with a delta pI = 0.04 pH units, have been selected. The load capacity increases almost linearly, as a function of progressively higher I values, from 0.5 X up to 2 X molarity of buffering Immobiline (pK 7.0) to abruptly reach a plateau at 3 X concentration of buffering ion. The load capacity also increases almost linearly as a function of gel thickness from 1 to 5 mm, without apparently levelling off. When decreasing the pH interval from 1 pH unit (pH 6.8-7.8) to 1/2 pH unit (pH 7.05-7.55) the amount of protein loaded in the HbA zone could be increased by 40%. In 5 mm thick gels, at 2 X pK 7.0 Immobiline concentration, over a 1/2 pH unit span, up to 350 mg HbA (in a 12.5 X 11 cm gel) could be loaded in a single zone, the load limit of the system being around 45 mg protein/ml gel volume. 相似文献
44.
E Gianazza G Dossi F Celentano P G Righetti 《Journal of biochemical and biophysical methods》1983,8(2):109-133
A new technique for generating extended pH gradients (5 pH units) in Immobiline gels is reported. The previously described (J. Biochem. Biophys. Methods 7, 1983, 123-142) five-chamber gradient mixer has been replaced by a two-vessel device. A single mixture of the available Immobilines (pK 3.6, 4.6, 6.2, 7.0, 8.5 and 9.3) is made, with relative concentrations adjusted so as to produce the most uniform buffering power throughout the desired pH interval. This mixture is then divided into two portions, which are titrated to the extremes of the required pH span with an acidic titrant (Immobiline pK approximately 1) and a basic species (Immobiline pK 9.95). Highly reproducible pH gradients (pH 4-9) are thus generated, which appear extremely useful for the first dimensioned of 2-dimensional techniques. Our previously reported computer program has been implemented with an optimization algorithm which, given any cocktail of Immobilines, automatically adjusts the relative initial concentrations until the smoothest possible beta power is found. For the first time it is possible to perform IEF under controlled physico-chemical parameters: pH span and linearity, beta power, ionic strength and molarity of the buffering species. 相似文献
45.
Microbial mats and physicochemistry in a saltern in the Bretagne (France) and in a laboratory scale saltern model 总被引:1,自引:0,他引:1
Dieter Giani Jacob Seeler Luise Giani Wolfgang E. Krumbein 《FEMS microbiology letters》1989,62(3):151-161
Abstract A saltern near La Baule (Bretagne, France) was remodeled in a programmable temperature and humidity controlled walk-in environmental chamber resembling the characteristics of the original saltern. The saltern showed different types of microbial mats predominantly composed of algae, oxy- and anoxyphotobacteria, and associated chemoorganotrophic bacteria, fungi and animals. Well-developed microbial mats were found up to a salinity of 10% during the three or four months in summer when salinity gradients and NaCl precipitation were established. The main phototrophic organisms were diatoms, the cyanobacteria Aphanothece, Microcoleus, Spirulina , and Oscillatoria , and Chromatiaceae. At higher salinity, Halobacterium sp., diatoms, and Dunaliella were dominant. Typical microbial mats and saltern-typical invertebrate, algal and bacterial species also developed in the saltern model, building up a stable community. The ionic composition of the brines and physicochemical parameters were similar to those determined for the original saltern. Different photosynthetic organisms, e.g. a filamentous purple bacterium and a hypersaline Chloroflexus -like organism, could be enriched within the microbial mats by changing the light regime. 相似文献
46.
The rapid transients of CO2 gas exchange have been measured in leaves ofHelianthus annuus L. In parallel experiments the assimilatory force FA, which is the product of the phosphorylation potential and the redox ratio NADPH/NADP, has been calculated from measured
ratios of dihydroxyacetone phosphate to phosphoglycerate in the chloroplast stroma and in leaves. The following results were
obtained: (i) When the light-dependent stroma alkalization was measured under steady-state conditions for photosynthesis in
air containing 2000 μl · l-1 CO2, alkalization increased with photosynthesis as the quantum flux density (irradiance) was increased. This contrasts to the
light-dependent stroma alkalisation measured in dark-adapted leaves during the dark-light transient (Laisk et al. 1989, Planta177, 350–358) which reached a maximum at a quantum flux density far below that necessary to saturate photosynthesis. This maximum
was about three times higher than the maximum stroma alkalization at light- and CO2-saturated photosynthesis. (ii) Accurate calculations of the assimilatory force FA require a consideration of the stromal pH. However, under many conditions, changes in the stromal pH resulting from changes
in photosynthetic flux can be neglected because they are small. (iii) Stromal ratios of dihydroxyacetone phosphate to phosphoglycerate
are generally lower than ratios measured in leaf extracts. The value of FA calculated from stromal metabolites was about 30% lower than FA calculated from cellular metabolites. Still, it appears sufficient for many purposes to calculate FA from metabolite measurements in leaf extracts. (iv) In the light, the catalytic capacity of the photosynthetic apparatus
is adjusted to the level of irradiance. The response of carbon assimilation to large increases in irradiance is slow because
it requires enzyme activation. Deactivation of the Calvin cycle induced by decreases in irradiance is slower than activation.
(v) Changes in catalytic capacity and in the availability or level of substrates such as CO2 alter the flux resistance of the Calvin cycle. A decrease in flux resistance explains why FA often does not increase by much and may actually decrease when carbon flux is increased. Adjustments of flux resistances
in the Calvin cycle and of photosystem-II activity in the electron-transport chain permit varying rates of photosynthesis
at low levels of ATP and NADPH. As NADP remains available, the danger of over-reduction which leads to photoinactivation of
electron transport is minimized.
K.R. und U.H. were guests of the Estonian Academy of Sciences. Support by the Estonian Academy of Sciences, the Sonderforschungsbereich
251 of the University of Würzburg and the Fonds der Chemischen Industrie is gratefully acknowledged. 相似文献
47.
Pranav K. Sinha E. K ttgen Marina St ffler-Meilicke Elisabetta Gianazza Pier Giorgio Righetti 《Journal of biochemical and biophysical methods》1990,20(4):345-352
Up to the present time it has been impossible to perform two-dimensional (2-D) separations in very acidic immobilized pH gradients (IPG), due to the lack of suitable buffering acrylamido derivatives to be incorporated into the polyacrylamide matrix. The advent of the pK 3.1 buffer (2-acrylamido glycolic acid; Righetti et al., J. Biochem. Biophys. Methods 16, 1988, 185–192) allowed the formulation of such acidic gradients. We report here separations in IPG pH 2.8–5.0 intervals of polypeptide chains from total lysates of rat intestinal and liver cells and 30S and 50S ribosomal proteins from Halobacterium marismortui. Conditions are given for highly reproducible first and second dimensions gels and for a proper silver staining of 2-D maps with practically no background deposition. 相似文献
48.
The present study is based on the hypothesis that nonuniform hemodynamics, represented by large time-averaged wall shear stress gradients, trigger abnormal biological processes leading to rapid restenosis, i.e. excessive tissue overgrowth and renewed plaque formation, and hence early graft failure. It implies that this problem may be significantly mitigated by finding graft-artery bypass configurations for which the wall shear stress gradient is approximately zero and hence nearly uniform hemodynamics is achieved. These fluid flow and geometric design considerations are applied to four different end-to-side anastomoses for the distal end of a femoral artery bypass with an appropriate test input pulse and a typical 20–80 flow division. A validated finite-volume code has been used to compute the transient three-dimensional velocity vector fields, wall shear stress distributions and surface contours of the wall shear stress gradients. It is shown that large anastomotic flow areas, small continuously changing bifurcation angles, and smooth junction wall curvatures reduce local time-averaged wall shear stress gradients significantly and hence should mitigate restenosis. 相似文献
49.
The interactions between phytoplankton and zooplankton were studied in two large lakes in the Saimaa lake system, Finland. Both are subjected to substantial waste water loading, and exhibit a clear trophic gradient between the loaded and unloaded areas. The phytoplankton and zooplankton were compared in terms of composition, abundance and biomass at 34–39 stations located in different parts of the lakes. At least four mechanisms were thought to affect the composition of plankton communities: (1) the amount of nutrients (trophic gradient), (2) grazing of algae by herbivores, (3) the effect of the algal species composition on feeding by zooplankters (large, colonial algae in the more loaded parts of the lakes) and (4) the regeneration and reorganization of nutrients. 相似文献
50.
Douglas A. Burns 《Biogeochemistry》1996,32(3):339-362
Calcite treatment of chronically acidic lakes has improved fish habitat, but the effects on downstream water quality have not previously been examined. In this study, the spatial and temporal effects of watershed CaCO3 treatment on the chemistry of a lake outlet stream in the Adirondack Mountains of New York were examined. Before CaCO3 treatment, the stream was chronically acidic. During spring snowmelt before treatment, pH and acid-neutralizing capacity (ANC) in the outlet stream declined, and NO
3
–
and inorganic monomeric aluminum (AlIM) concentrations increased sharply. During that summer, SO
4
–
and NO
3
–
concentrations decreased downstream, and dissolved organic carbon (DOC) concentrations and ANC increased, in association with the seasonal increase in decomposition of organic matter and the attendant SO
4
–
-reduction process. A charge-balance ANC calculation closely matched measured downstream changes in ANC in the summer and indicated that SO
4
–
reduction was the major process contributing to summer increases in ANC. Increases in Ca2+ concentration and ANC began immediately after CaCO3 application, and within 3 months, exceeded their pretreatment values by more than 130 eq/L. Within 2 months after treatment, downstream decreases in Ca2+ concentration, ANC, and pH, were noted. Stream mass balances between the lake and the sampling site 1.5 km downstream revealed that the transport of all chemical constituents was dominated by conservative mixing with tributaries and ground water; however, non-conservative processes resulted in significant Ca2+ losses during the 13-month period after CaCO3 treatment. Comparison of substrate samples from the buffered outlet stream with those from its untreated tributaries showed that the percentage of cation-exchange sites occupied by Ca2+ as well as non-exchangeable Ca, were higher in the outlet-stream substrate than in tributary-stream substrate. Mass-balance data for Ca2+ H+, AlIM, and DOC revealed net downstream losses of these constituents and indicated that a reasonable set of hypothesized reactions involving AlIM, HCO
3
–
, Ca2+, SO
4
–
NO
3
–
, and DOC could have caused the measured changes in stream acid/base chemistry. In the summer, the sharp decrease in ANC continued despite significant downstream decreases in SO4
2– concentrations. After CaCO3 treatment, reduction of SO
4
–
was only a minor contributor to ANC changes relative to those caused by Ca2+ dilution from acidic tributaries and acidic ground water, and Ca2+ interactions with stream substrate. 相似文献