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1.
The effect of toluene on the structure and permeability of the outer and cytoplasmic membranes of Escherichia coli. 总被引:8,自引:0,他引:8
The effect of toluene on Escherichia coli has been examined. In the presence of Mg2+, toluene removes very little protein, phospholipid, or lipopolysacharide from E. coli. In the absence of Mg2+, or in the presence of EDTA, toluene removes considerably more cell material, including several specific cytoplasmic proteins such as malate dehydrogenase (EC 1.1.1.37). In contrast, glucose-6-phosphate dehydrogenase (EC 1.1.1.49) and glutamate dehydrogenase (EC 1.4.1.4) are not released at all under the same conditions. Cells treated with toluene in the presence of Mg2+ remain relatively impermeable to pyridne nucleotides, while cells treated with toluene in the presence of EDTA become permeable to these compounds. Freeze-fracture electron microscopy shows that toluene causes considerable damage to the cytoplasmic membrane, while the outer membrane remains relatively intact. These results indicate that the permeability characteristics of toluene-treated cells depend at least partly on the state of the outer membrane after the toluene treatment. 相似文献
2.
Lysozyme fails to penetrate through the outer membrane of stationary phase cells of Escherichia coli when it is simply added to suspensions of plasmolyzed cells. Lysozyme penetrates the outer membrane only when these cells are exposed to a mild osmotic shock in the presence of EDTA and lysozyme.In the presence of Mg2+, the outer membrane is stabilized sufficiently so that there is no lysozyme penetration during osmotic shock. If Mg2+ is added after an osmotic shock has been used to cause lysozyme to penetrate a destabilized outer membrane, the outer membrane is stabilized once again. In this case however, cells are converted to spheroplasts by the lysozyme which has gained access to the murein layer prior to the addition of Mg2+. Mg2+ stabilizes the outer membranes of these spheroplasts sufficiently so that they remain immune to lysis even in the absence of osmotic stabilizers such as sucrose.These results are discussed in terms of current information on the structure of the murein layer and the outer membrane. 相似文献
3.
Function of Teichoic Acids and Effect of Novobiocin on Control of Mg<Superscript>2+</Superscript> at the Bacterial Membrane 总被引:11,自引:0,他引:11
ALTHOUGH the occurrence of both wall and membrane teichoic acids in Gram-positive bacteria has been known for a considerable time and it is believed that they are essential for normal cellular activity, their main function has been somewhat obscure. Confirmatory evidence for the proposal1 that teichoic acids participate in ion-exchange in the outer regions of the bacterial cell has been described recently2. It has been shown that the phosphate groups of the wall teichoic acid are responsible for the capacity of isolated walls to bind magnesium ions; but whole cells of Gram-positive bacteria also invariably contain a poly-glycerol phosphate-teichoic acid located in the region between the wall and the cytoplasmic membrane3 and it is believed that this must be able to bind Mg2+ as does the wall polymer. These two regions of anionic polymer might thus constitute an integrated cation-exchange system between the exterior of the cell and the cytoplasmic membrane, where relatively high concentrations of Mg2+ are required for a variety of processes. We report here experiments with a membrane-bound enzyme system that requires Mg2+, obtained from a broken cell preparation and in which the close contact between the outer layers of the cell is preserved. In this preparation the enzyme system displays maximum activity in the presence of Mg2+ bound to the endogenous teichoic acid and is insensitive to changes in the concentration of added Mg2+, in marked contrast to the behaviour of the enzyme system in isolated cytoplasmic membrane. These results provide the first direct demonstration of the function of teichoic acids in concentrating Mg2+at the cytoplasmic membrane. They lead to the conclusion that failure of teichoic acid biosynthesis in the whole cell would cause inhibition of membrane function through magnesium starvation. In view of this the effect of novobiocin, an antibiotic shown to inhibit teichoic acid biosynthesis in vitro4–6, is discussed. 相似文献
4.
Parameters involved in reconstitution of the outer membrane permeability described by Brunner, Caputo, and Treick [3] were
examined. The most efficient reconstitution was obtained when divalent cations accompanied the addition of exogenous outer
membrane material. Studies indicated that the effectiveness of Ca2+ or Mg2+ to promote reassociation of outer membrane material, and subsequent protection against actinomycin D, was dependent upon
the strain ofEscherichia coli. More specifically, the data suggest that the effectiveness of the different divalent cations in promoting reassociation
was determined by the relative amounts of F1 and F2 fractions released by ethylenediaminetetraacetate (EDTA). Reconstitution
was shown by cell survival to be as high as 25% and dependent upon the total amount of material released from the outer membrane
by EDTA. Between 50 and 80% of the bound material could be removed from the cells by subsequent EDTA treatment. 相似文献
5.
Shigehiko Miyamoto 《生物化学与生物物理学报:生物膜》2007,1768(7):1848-1854
The LolCDE complex of Escherichia coli releases outer membrane-specific lipoproteins from the inner membrane. Lipoproteins with Asp at + 2 remain in the inner membrane since this residue functions as a LolCDE avoidance signal depending on phosphatidylethanolamine. We examined the effects of other phospholipids on lipoprotein sorting in proteoliposomes reconstituted with LolCDE and various synthetic phospholipids. The lipoprotein release and ATP hydrolysis were both low at 2 mM Mg2+ but very high at 10 mM Mg2+ in proteoliposomes containing cardiolipin alone. However, the Lol avoidance function was abolished at 10 mM Mg2+, and the release of lipoproteins with Asp at + 2 was as efficient as that of outer membrane-specific lipoproteins. The addition of phosphatidylethanolamine to cardiolipin stimulated the ATP hydrolysis and increased the Lol avoidance function of Asp at + 2 at 2 mM Mg2+. The addition of phosphatidylglycerol to cardiolipin nearly completely inhibited the release of lipoproteins with Asp at + 2 even at 10 mM Mg2+, while that of outer membrane-specific lipoproteins was not. Taken together, these results indicate that three major phospholipids of E. coli differently affect lipoprotein sorting and the activity of LolCDE. 相似文献
6.
Interactions of Cations with Membrane Fractions of Smooth and Rough Strains of Salmonella typhimurium and Other Gram-Negative Bacteria 总被引:1,自引:1,他引:0
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Addition of cations (20 to 50 mM for Mg2+ or Ca2+ or 100 to 500 mM for Na+) to N-2-hydroxyethylpiperazine-N′-2-ethanesulfonic acid buffer during preparation of membranes from smooth and rough strains of Salmonella typhimurium LT2, Salmonella minnesota, and Escherichia coli O8 had two effects on the composition of the membranes isolated. First, in rough strains of chemotypes Ra to Re the “total membranes” (pellets from high-speed centrifugation) were deficient in the proteins of the outer membrane. The missing proteins were found to have been sedimented in a prior low-speed centrifugation in a fraction we call “cation-aggregated membranes.” Since these membranes were enriched for lipopolysaccharide and for outer membrane proteins, deficient in succinic dehydrogenase, and contained primarily the dense peak after sucrose gradient centrifugation, it appears to be relatively pure outer membrane. About 10% of the membrane protein of smooth strains and up to 50% that of rough strains were cation-aggregated membranes, appearing to contain most of the outer membrane of rough strains. Thus, cation aggregation may be a useful means of preparation of outer membrane samples. The second effect was that with cation addition, several high-molecular-weight proteins not seen when membranes were prepared without cation addition were found in the total membranes of both smooth and rough strains after high-speed centrifugation. These proteins were bound by cations to the inner membranes, since they were soluble in Triton X-100 and separated into the less dense peak upon sucrose gradient centrifugation. They originated from the cytoplasm or the periplasm, since they corresponded to soluble proteins found in the supernatant after high-speed centrifugation and were depleted from this supernatant when preparation was done in the presence of cations. 相似文献
7.
Reconstitution of the energy-linked transhydrogenase activity in membranes from a mutant strain of Escherichia coli K12 lacking magnesium ion- or calcium ion-stimulated adenosine triphosphatase 总被引:9,自引:9,他引:0
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G. B. Cox F. Gibson L. M. McCann J. D. Butlin F. L. Crane 《The Biochemical journal》1973,132(4):689-695
1. We have isolated a mutant of Escherichia coli K12 (strain AN295) that forms de-repressed amounts of Mg2+,Ca2+-stimulated adenosine triphosphatase. 2. The Mg2+,Ca2+-stimulated triphosphatase activity was separated from membrane preparations from strain AN295 by extraction with 5mm-Tris–HCl buffer containing EDTA and dithiothreitol, resulting in a loss of the ATP-dependent transhydrogenase activity. The non-energy-linked transhydrogenase activity remained in the membrane residue. 3. The solubilized Mg2+,Ca2+-stimulated adenosine triphosphatase activity from strain AN295 was partially purified by repeated gel filtration. The addition of the purified Mg2+,Ca2+-stimulated adenosine triphosphatase to the membrane residue from strain AN295 reactivated the ATP-dependent transhydrogenase activity. 4. Strain AN296, lacking Mg2+,Ca2+-stimulated adenosine triphosphatase activity, was derived by transducing the mutant allele, uncA401, into strain AN295. The ATP-dependent transhydrogenase activity was lost but the non-energy linked transhydrogenase was retained. 5. The ATP-dependent transhydrogenase activity in membrane preparations from strain AN296 (uncA−) could not be re-activated by the purified Mg2+,Ca2+-stimulated adenosine triphosphatase from strain AN295. However, after extraction by 5mm-Tris–HCl buffer containing EDTA and dithiothreitol, the ATP-dependent transhydrogenase activity could be re-activated by the addition of the purified Mg2+,Ca2+-stimulated adenosine triphosphatase from strain AN295 to the membrane residue from strain AN296 (uncA−). 相似文献
8.
Uptake and degradation of EDTA by Escherichia coli 总被引:1,自引:0,他引:1
It was found that Escherichia coli exhibited a growth by utilization of Fe(III)EDTA as a sole nitrogen source. No significant growth was detected when Fe(III)EDTA
was replaced by EDTA complexes with other metal ions such as Ca2+, Co2+, Cu2+, Mg2+, Mn2+, and Zn2+. When EDTA uptake was measured in the presence of various ions, it was remarkable only when Fe3+ was present. The cell extract of E. coli exhibited a significant degradation of EDTA only in the presence of Fe3+. It is likely that the capability of E. coli for the growth by utilization of Fe(III)EDTA results from the Fe3+-dependent uptake and degradation of EDTA. 相似文献
9.
A method was developed for the reassembly of membranous vesicle from the sodium dcoxycholate-dissociated outer membrane components of Escherichia coli. The removal of the detergent by dialysis and the presence of Mg2+ were essential for the reassembly.Membrane protein alone did not form any membranous structure. Closed membranous vesicles similar to the native outer membrane were reassembled only when protein was mixed with both lipopolysaccharide and phospholipid in deoxycholate solution and subsequently dialyzed. The membrane showed a distinct trilaminar structure with a center-to-center distance between two dark lines of 53 Å, which is a characteristic of the native outer membrane. This characteristic trilaminar structure was shown to be due to the presence of lipopolysaccharide. Phospholipd was required for the vesicularization of membrane. Lipopolysaccharide and/or phospholipid formed a membranous structure in the absence of protein, while the morphology of their negatively stained sample was quite different from that of the native outer membrane unless the outer membrane protein was added to the reassembly mixture.The protein from the cytoplasmic membrane was unable to reform membranous vesicle with lipopolysaccharide and phospholipid, indicating that the reassembly system discriminated outer membrane proteins from cytoplasmic membrane proteins. 相似文献
10.
Ultrastructural and Enzymic Studies of Cell Membranes from Ice-encased and Noniced Winter Wheat Seedlings
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A marked increase in the amount of cisternal-like cytoplasmic membranes was observed after ice encasement of winter wheat (Triticum aestivum L.) seedlings. Linear sucrose gradients were employed to separate the various membrane components of the microsomal membrane fraction. NADH- and NADPH-cytochrome c reductase, two specific enzyme markers for plant endoplasmic reticulum (ER) were used to locate the ER in the linear gradients. The identity of the ER fraction was confirmed by determining the effect of EDTA and Mg2+ in the preparative media on the distribution of NADH- and NADPH-cytochrome c reductase activity within the gradient. In the presence of EDTA which dissociates ribosomes from ER, peaks of activity for the two enzymes were observed at a density corresponding to that for “smooth” ER. When the media also contained an appropriate concentration of Mg2+ to maintain the attachment of ribosomes to the ER, the peaks of activity for the enzymes shifted to a density corresponding to that for “rough” ER. NADH-cytochrome c reductase activity was similar for 24 C-grown and 2 C-grown iced seedlings, but significantly lower for 2 C noniced seedlings. No preferential increase in uptake of radioactive leucine or choline in the ER was observed during ice encasement. The accumulation of electron microscopically visible membrane arrays was not inhibited by the presence of protein synthesis inhibitors at concentrations which severely inhibited incorporation of [1-14C]leucine into membrane protein, but did not affect survival and growth of the seedlings. These observations indicate that the apparent proliferation of ER during ice encasement does not result from net membrane synthesis, but rather from reorganization of existing membrane elements within the cell. 相似文献
11.
The formation of the first intermediate in ubiquinone-9 biosynthesis, 3-nonaprenyl-4-hydroxybenzoate (NPHB), by the enzyme p-hydroxybenzoate:polyprenyl transferase, has been studied in isolated rat liver mitochondria using solanesol pyrophosphate and p-hydroxybenzoate as the substrates. Phosphate buffer (100 mm) is inhibitory but at 20 mm inhibition is not apparent compared to other buffers at the same concentration. With various buffers at low concentration (20 mm) both EDTA and Mg2+ stimulate formation of NPHB while Ca2+ inhibits. Release of Ca2+ inhibition can be achieved by the addition of Mg2+, or EDTA, or EGTA, with EGTA being less effective than EDTA. When Mg2+, Ca2+, and EDTA are present together, a two- to threefold increase in activity of the enzyme is observed. The antibiotic bacitracin inhibits the synthesis of NPHB and the inhibition is increased when divalent cations are present. EGTA is more effective than EDTA in overcoming inhibition due to bacitracin. The possibility that these effects are partially due to alteration of mitochondrial membrane conformation as well as a direct effect on the enzyme is evaluated. The possible role of polyprenylphosphates in mitochondrial membrane function is discussed. 相似文献
12.
We have investigated the possible relationships between the cation-induced and phenazine methosulfate (PMS)-induced fluorescence changes and their relation to light induced conformational changes of the thylakoid membrane.1. In isolated chloroplasts, PMS markedly lowers the quantum yield of chlorophyll a fluorescence (φf) when added either in the presence or the absence of dichloro-phenyldimethylurea (DCMU). In contrast, Mg2+ causes an increase in φf. However, these effects are absent in isolated chloroplasts fixed with glutaraldehyde that retain (to a large extent) the ability to pump protons, suggesting that structural alteration of the membrane—not the pH changes—is required for the observed changes in φf. The PMS triggered decrease in φf is not accompanied by any changes in the emission (spectral) characteristics of the two pigment systems, whereas room temperature emission spectra with Mg2+ and Ca2+ show that there is a relative increase of System II to System I fluorescence.2. Washing isolated chloroplasts with 0.75 mM EDTA eliminates (to a large extent) the PMS-induced quenching and Mg2+-induced increase of φf, and these effects are not recovered by the further addition of dicyclohexyl carbodiimide. It is known that washing with EDTA removes the coupling factor, and thus, it seems that the coupling factor is (indirectly) involved in conformational change of thylakoid membranes leading to fluorescence yield changes.3. In purified pigment System II particles, neither PMS nor Mg2+ causes any change in φf. Our data, taken together with those of the others, suggest that a structural modification of the thylakoid membranes (not macroscopic volume changes of the chloroplasts) containing both Photosystems I and II is necessary for the PMS-induced quenching and Mg2+-induced increase of φf. These two effects can be explained with the assumption that the PMS effect is due to an increase in the rate of internal conversion (kh), whereas the Mg2+ effect is due to a decrease in the rate of energy transfer (kt), between the two photosystems.4. From the relative ratio of φf with DCMU and DCMU plus Mg2+, we have calculated kt (the rate constant of energy transfer between Photosystems II and I to be 4.2·108 s?1, and φt (quantum yield of this transfer) to be 0.12. 相似文献
13.
Membrane Damage and Microbial Inactivation by Chlorine in the Absence and Presence of a Chlorine-Demanding Substrate 总被引:1,自引:0,他引:1
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The relationship between cell inactivation and membrane damage was studied in two gram-positive organisms, Listeria monocytogenes and Bacillus subtilis, and two gram-negative organisms, Yersinia enterocolitica and Escherichia coli, exposed to chlorine in the absence and presence of 150 ppm of organic matter (Trypticase soy broth). L. monocytogenes and B. subtilis were more resistant to chlorine in distilled water. The addition of small amounts of organic matter to the chlorination medium drastically increased the resistance of both types of microorganisms, but this effect was more marked in Y. enterocolitica and E. coli. In addition, the survival curves for these microorganisms in the presence of organic matter had a prolonged shoulder. Sublethal injury was not detected under most experimental conditions, and only gram-positive cells treated in distilled water showed a relevant degree of injury. The exposure of bacterial cells to chlorine in distilled water caused extensive permeabilization of the cytoplasmic membrane, but the concentrations required were much higher than those needed to inactivate cells. Therefore, there was no relationship between the occurrence of membrane permeabilization and cell death. The addition of organic matter to the treatment medium stabilized the cytoplasmic membrane against permeabilization in both the gram-positive and gram-negative bacteria investigated. Exposure of E. coli cells to the outer membrane-permeabilizing agent EDTA increased their sensitivity to chlorine and caused the shoulders in the survival curves to disappear. Based on these observations, we propose that bacterial envelopes could play a role in cell inactivation by modulating the access of chlorine to the key targets within the cell. 相似文献
14.
Strain NR70, a mutant of lacking the Mg2+-adenosine triphosphatase (E.C. 3.6.1.3.) was previously shown to be defective in amino acid and sugar transport in whole cells and right-side-out membrane vesicles. It is shown here that the mutant is also deficient in the uptake of calcium into inverted membrane vesicles. Treatment of inverted vesicles from the wild-type strain with ethylenediamine tetraacetate removes the Mg2+-adenosine triphosphatase and results in an inability to transport calcium. Addition of a crude fraction containing the wild-type Mg2+-adenosine triphosphatase restores active uptake of calcium both to vesicles from the mutant and depleted vesicles from the wild-type. 相似文献
15.
Kojiro Takahashi Henry Weiner James H.J. Hu 《Archives of biochemistry and biophysics》1980,205(2):571-578
The V of horse liver aldehyde dehydrogenase is enhanced twofold in the presence of 0.5 mm Mg2+ ions when assayed in the dehydrogenase reaction. The mechanism of this activation appears to be related to the fact the enzyme changes from functioning with half-of-the-sites reactivity to functioning with all-of-the-sites reactivity. That is, the presteady-state burst magnitude increases from 2 mol NADH formed per mole of tetrameric enzyme to 4 mol formed per mole (K. Takahashi and H. Weiner, J. Biol. Chem., 1980, 255, 8206–8209). Whether this twofold enhancement correlates, in fact, to a change from half-of-the-sites to all-of-the-sites reactivity of the enzyme by Mg2+ ions was investigated by determining the Stoichiometry of coenzyme binding by fluorescence quenching and enhancement methods in the absence and presence of the metal ions. The biphasic Scatchard plots for NAD binding to the enzyme were similar in the absence and presence of Mg2+ ions, while that of NADH binding was monophasic (-Mg2+) and biphasic (+Mg2+). In the presence of p-methoxyacetophenone, a competitive inhibitor for substrate, the stoichiometric titration of coenzyme binding to the ternary complexes (enzyme-NAD(H)-inhibitor) revealed that only 2 mol of NAD or NADH bind in the absence of Mg2+ ions but 4 bind per mole of tetrameric enzyme in the presence of added metal. The fluorescence intensity of NAD's fluorescent derivative, 1,N6-ethenoadenine dinucleotide, bound to the enzyme was also doubled by the addition of Mg2+ ions.The combined binding data show that the stoichiometry of coenzyme binding to aldehyde dehydrogenase in the ternary complex increases from 2 to 4 mol binding per mole of tetrameric enzyme with the addition of Mg2+ ions. This increase in stoichiometry corresponds to the observed changes of burst magnitude obtained from the presteady-state and V in the steady-state kinetics assays. From both results of the kinetics and stoichiometry, we show that horse liver aldehyde dehydrogenase exhibits half-of-the-sites reactivity when in the tetrameric state in the absence of Mg2+ ions, and all-of-the-sites reactivity in the dimeric state in the presence of the metal. 相似文献
16.
Kafa Walweel Jiao Li Peter Molenaar Mohammad S. Imtiaz Anthony Quail Cris G. dos Remedios Nicole A. Beard Angela F. Dulhunty Dirk F. van Helden Derek R. Laver 《The Journal of general physiology》2014,144(3):263-271
Regulation of the cardiac ryanodine receptor (RyR2) by intracellular Ca2+ and Mg2+ plays a key role in determining cardiac contraction and rhythmicity, but their role in regulating the human RyR2 remains poorly defined. The Ca2+- and Mg2+-dependent regulation of human RyR2 was recorded in artificial lipid bilayers in the presence of 2 mM ATP and compared with that in two commonly used animal models for RyR2 function (rat and sheep). Human RyR2 displayed cytoplasmic Ca2+ activation (Ka = 4 µM) and inhibition by cytoplasmic Mg2+ (Ki = 10 µM at 100 nM Ca2+) that was similar to RyR2 from rat and sheep obtained under the same experimental conditions. However, in the presence of 0.1 mM Ca2+, RyR2s from human were 3.5-fold less sensitive to cytoplasmic Mg2+ inhibition than those from sheep and rat. The Ka values for luminal Ca2+ activation were similar in the three species (35 µM for human, 12 µM for sheep, and 10 µM for rat). From the relationship between open probability and luminal [Ca2+], the peak open probability for the human RyR2 was approximately the same as that for sheep, and both were ∼10-fold greater than that for rat RyR2. Human RyR2 also showed the same sensitivity to luminal Mg2+ as that from sheep, whereas rat RyR2 was 10-fold more sensitive. In all species, modulation of RyR2 gating by luminal Ca2+ and Mg2+ only occurred when cytoplasmic [Ca2+] was <3 µM. The activation response of RyR2 to luminal and cytoplasmic Ca2+ was strongly dependent on the Mg2+ concentration. Addition of physiological levels (1 mM) of Mg2+ raised the Ka for cytoplasmic Ca2+ to 30 µM (human and sheep) or 90 µM (rat) and raised the Ka for luminal Ca2+ to ∼1 mM in all species. This is the first report of the regulation by Ca2+ and Mg2+ of native RyR2 receptor activity from healthy human hearts. 相似文献
17.
Kazuei Igarashi Kazuhiro Mitsui Masae Kubota Mari Shirakuma Reiko Ohnishi Seiyu Hirose 《Biochimica et Biophysica Acta (BBA)/General Subjects》1983,755(3):326-331
The effect of polyamines on the in vitro and in vivo synthesis and degradation on guanosine 5′-diphosphate 3′-diphosphate (ppGpp) has been studied in Escherichia coli. The presence of 2 mM spermidine lowered the optimal Mg2+ concentration for ppGpp formation from 17 mM to 11 mM. The formation of ppGpp in the presence of 2 mM spermidine and 11 mM Mg2+ was about 15% greater than that in the presence of 17 mM Mg2+. At a concentration of less than 11 mM Mg2+, spermidine was found to stimulate ppGpp formation greatly. Putrescine did not cause any effect. When a polyamine-requiring mutant of E. coli (EWH319) was starved for an amino acid by the addition of valine, spermidine stimulated ppGpp formation. the degradation of ppGpp was not influenced significantly by polyamines. 相似文献
18.
A cell envelope fraction has been prepared after mechanical disruption of lysozyme-EDTA spheroplasts from depigmented Rhodopseudomonas sphaeroides (aerobically grown in the light). On linear sucrose gradients this fraction can be separated in a cytoplasmic membrane fraction and an outer membrane fraction. The cytoplasmic fraction (buoyant density: 1.18 g/cm3) has been characterized by its succinic dehydrogenase activity and by its composition. The outer membrane fraction (buoyant density: 1.21 g/cm3) does not contain any respiratory activity nor hemoproteins. The same fractionation has been done on cells repigmented in the dark by lowering the O2 pressure. In that case the same two fractions have been detected in addition to the chromatophore fraction (buoyant density: 1.14 g/cm3). However both, and specially the outer membrane fraction, were contaminated by chromatophore material. 相似文献
19.
Because mitochondrial dysfunction occurs relatively early in the course of nephrotoxicity associated with the aminoglycoside antibiotic, gentamicin, the acute in vitro effects of gentamicin on renal cortical mitochondrial respiration were studied. Gentamicin produced stimulation of State 4 rates and inhibition of State 3 and DNP-uncoupled rates with pyruvate-malate or succinate as substrates. The stimulation of State 4 respiration was not blocked by oligomycin. Both the stimulation of State 4 and inhibition of State 3 were profoundly dependent on the Na+ and K+ contents of the incubation medium, were potentiated by the presence of EDTA, and were reversed by Mg2+. These results suggested that gentamicin's effects on mitochondrial respiration were due to alterations in the interaction of Na+ and K+ with the inner mitochondrial membrane at Mg2+-sensitive sites. 相似文献
20.
The uptake of proline and glutamine by cytochrome-deficient cells of Escherichia coli SASX76 grown aerobically on glucose or anaerobically on pyruvate was stimulated by these two substrates. Pyruvate could not stimulate transport in the glucose-grown cells. Uptake of these amino acids energized by glucose was inhibited by inhibitors of the Ca2+, Mg2+-stimulated ATPase such as DCCD, pyrophosphate, and azide, and by the uncouplers CCCP and 2,4-dinitrophenol. Glycerol (or glycerol 3-phosphate) in the presence of fumarate stimulated the transport of proline and glutamine under anaerobic conditions in cytochrome-deficient cells but not in membrane vesicles prepared from these cells although glycerol 3-phosphate-fumarate oxidoreductase activity could be demonstrated in the vesicle preparation. In contrast, in vesicles prepared from cytochrome-containing cells of E. coli SASX76 amino acid transport was energized under anaerobic conditions by this system. Inhibitors of the Ca2+, Mg2+-activated ATPase and uncoupling agents inhibited the uptake of proline and glutamine in cytochrome-deficient cells dependent on the glycerol-fumarate oxidoreductase system. Ferricyanide could replace fumarate as an electron acceptor to permit transport of phenylalanine in cytochrome-deficient or cytochrome- containing cells under anaerobic conditions. It is concluded that in cytochrome-deficient cells using glucose, pyruvate, or glycerol in the presence of fumarate, transport of both proline and glutamine under anaerobic conditions is energized by ATP through the Ca2+, Mg2+-activated ATPase. In cytochrome-containing cells under anaerobic conditions electron transfer between glycerol and fumarate can also drive transport of these amino acids. 相似文献