共查询到20条相似文献,搜索用时 31 毫秒
1.
Osmotic swelling of fish erythrocytes activates a broad-specificity permeation pathway that mediates the volume-regulatory
efflux of taurine and other intracellular osmolytes. This pathway is blocked by inhibitors of the erythrocyte band 3 anion
exchanger, raising the possibility that band 3 is involved in the volume-regulatory response. In this study of eel erythrocytes,
a quantitative comparison of the pharmacology of swelling-activated taurine transport with that of band 3-mediated SO2−
4 transport showed there to be significant differences between them. N-ethylmaleimide and quinine were effective inhibitors of swelling-activated taurine transport but caused little, if any, inhibition
of band 3. Conversely, DIDS was a more potent inhibitor of band 3-mediated SO2−
4 flux than of swelling-activated taurine transport. In cells in isotonic medium, pretreated then co-incubated with 0.1 mm DIDS, the band 3-mediated transport of SO2−
4 and Cl− was reduced to a low level. Exposure of these cells to a hypotonic medium containing 0.1 mm DIDS was followed by the activation of a Cl− permeation pathway showing the same inhibitor sensitivity as swelling-activated taurine transport. The data are consistent
with swelling-activated transport of taurine and Cl− being via a common pathway. A comparison of the swelling-activated transport rates for taurine and Cl− with those for several other solutes was consistent with the hypothesis that this pathway is an anion-selective channel,
similar to those that mediate the volume-regulatory efflux of Cl− and organic osmolytes from mammalian cells.
Received: 7 July 1995/Revised: 2 September 1995 相似文献
2.
Carbazole (CA) is a heterocyclic nitrogen compound contained in the crude petroleum oil and recalcitrant to removal through
the refinery processes. For development of the efficient CA-degradation bioprocess, conditions for the recycle use of Sphingomonas sp. CDH-7 resting cells were examined. When the resting cells (O.D.660 3.3) were shaken in 50 mM K2HPO4-KH2PO4 buffer (pH 7.0) containing CA 1000 mg/L, CA 880 mg/L was degraded within 3 h, but thereafter the activity decreased markedly.
However, the activity was found to be restored to the initial level after the shaking treatment for 3 h in CA-free medium
solution or in the buffer containing 20 mM MgCl2. Although the CA-degradation activity of CDH-7 resting cells was lost after 3 h of shaking in the buffer containing 100 mM EDTA, it was restored through the shaking treatment for 3 h in the buffer containing 20 mM MgCl2. When CA was periodically added eight times at a concentration of 100 mg/L (0.599 mM) to the reaction mixture containing the resting cells, CA 778 mg/L (4.66 mM) was continuously degraded within 35 h by the recycle use of resting cells, with the restoration treatment after each CA-degradation
reaction by the resting cells.
Received: 28 June 2001 / Accepted: 30 July 2001 相似文献
3.
Characterization of Regulatory Volume Behavior by Fluorescence Quenching in Human Corneal Epithelial Cells 总被引:2,自引:0,他引:2
An in-depth understanding of the mechanisms underlying regulatory volume behavior in corneal epithelial cells has been in
part hampered by the lack of adequate methodology for characterizing this phenomenon. Accordingly, we developed a novel approach
to characterize time-dependent changes in relative cell volume induced by anisosmotic challenges in calcein-loaded SV40-immortalized
human corneal epithelial (HCE) cells with a fluorescence microplate analyzer. During a hypertonic challenge, cells shrank
rapidly, followed by a temperature-dependent regulatory volume increase (RVI), τc = 19 min. In contrast, a hypotonic challenge induced a rapid (τc = 2.5 min) regulatory volume decrease (RVD). Temperature decline from 37 to 24°C reduced RVI by 59%, but did not affect RVD.
Bumetanide (50 μM), ouabain (1 mM), DIDS (1 mM), EIPA (100 μM), or Na+-free solution reduced the RVI by 60, 61, 39, 32, and 69%, respectively. K+, Cl− channel and K+-Cl− cotransporter (KCC) inhibition obtained with either 4-AP (1 mM), DIDS (1 mM), DIOA (100 μM), high K+ (20 mM) or Cl−-free solution, suppressed RVD by 42, 47, 34, 52 and 58%, respectively. KCC activity also affects steady-state cell volume,
since its inhibition or stimulation induced relative volume alterations under isotonic conditions. Taken together, K+ and Cl− channels in parallel with KCC activity are important mediators of RVD, whereas RVI is temperature-dependent and is essentially
mediated by the Na+-K+-2Cl− cotransporter (Na+-K+-2Cl−) and the Na+-K+ pump. Inhibition of K+ and Cl− channels and KCC but not Na+-K+-2Cl− affect steady-state cell volume under isotonic conditions. This is the first report that KCC activity is required for HCE
cell volume regulation and maintenance of steady-state cell volume. 相似文献
4.
Solubilization of enzymes in water-in-oil microemulsions and their rapid and efficient release through use of a pH-degradable surfactant 总被引:3,自引:0,他引:3
α-Chymotrypsin and lysozyme were solubilized in a water/O-[(2-tridecyl, 2-ethyl-1,3-dioxolan-4-yl)methoxy]–O′-methoxy poly(ethylene glycol) (CK-2,13 surfactant)/isooctane water-in-oil microemulsion solution at 1.5–2 and 10 g l−1 for 0.15 and 1.2 M CK-2,13, respectively. Upon contact with an equal volume of 0.1 M NaH2PO4/Na2HPO4 buffer, pH 5, a three-phase system (Winsor-III system) was formed, consisting of a surfactant-rich middle phase and aqueous
and isooctane-rich “excess” phases. Both enzymes were rapidly released into the aqueous excess phase, with 70% recovery of
each in 30 and 60 min for microemulsion solutions containing 0.15 and 1.2 M surfactant, respectively. The recovered enzymes retained >90% of their original specific activity. 相似文献
5.
In frog red blood cells, K-Cl cotransport (i.e., the difference between ouabain-resistant K fluxes in Cl and NO3) has been shown to mediate a large fraction of the total K+ transport. In the present study, Cl−-dependent and Cl−-independent K+ fluxes via frog erythrocyte membranes were investigated as a function of external and internal K+ ([K+]
e
and [K+]
i
) concentration. The dependence of ouabain-resistant Cl−-dependent K+ (86Rb) influx on [K+]
e
over the range 0–20 mm fitted the Michaelis-Menten equation, with an apparent affinity (K
m
) of 8.2 ± 1.3 mm and maximal velocity (V
max
) of 10.4 ± 1.6 mmol/l cells/hr under isotonic conditions. Hypotonic stimulation of the Cl−-dependent K+ influx increased both K
m
(12.8 ± 1.7 mm, P < 0.05) and V
max
(20.2 ± 2.9 mmol/l/hr, P < 0.001). Raising [K+]
e
above 20 mm in isotonic media significantly reduced the Cl−-dependent K+ influx due to a reciprocal decrease of the external Na+ ([Na+]
e
) concentration below 50 mm. Replacing [Na+]
e
by NMDG+ markedly decreased V
max
(3.2 ± 0.7 mmol/l/hr, P < 0.001) and increased K
m
(15.7 ± 2.1 mm, P < 0.03) of Cl−-dependent K+ influx. Moreover, NMDG+ Cl substitution for NaCl in isotonic and hypotonic media containing 10 mm RbCl significantly reduced both Rb+ uptake and K+ loss from red cells. Cell swelling did not affect the Na+-dependent changes in Rb+ uptake and K+ loss. In a nominally K+(Rb+)-free medium, net K+ loss was reduced after lowering [Na+]
e
below 50 mm. These results indicate that over 50 mm [Na+]
e
is required for complete activation of the K-Cl cotransporter. In nystatin-pretreated cells with various intracellular K+, Cl−-dependent K+ loss in K+-free media was a linear function of [K+]
i
, with a rate constant of 0.11 ± 0.01 and 0.18 ± 0.008 hr−1 (P < 0.001) in isotonic and hypotonic media, respectively. Thus K-Cl cotransport in frog erythrocytes exhibits a strong asymmetry
with respect to transported K+ ions. The residual, ouabain-resistant K+ fluxes in NO3 were only 5–10% of the total and were well fitted to linear regressions. The rate constants for the residual influxes were
not different from those for K+ effluxes in isotonic (∼0.014 hr−1) and hypotonic (∼0.022 hr−1) media, but cell swelling resulted in a significant increase in the rate constants.
Received: 19 November 1998/Revised: 23 August 1999 相似文献
6.
W. W. Yotis 《Antonie van Leeuwenhoek》1965,31(1):84-94
Two to 4 mg/ml of an antibacterial agent occurring in the serum of humans and animals caused the clotting of citrated rabbit, human, calf, ox, sheep, goat, guinea pig, rat, mouse, horse, chicken, pigeon and swine plasma. Heparinized plasmas of the same species were found resistant to the clotting action of the antibacterial agent. Citrated plasma previously heated at 56 C for 30 min, treated with 640 units/ml tyrosinase for 60 min, or absorbed with 0.2M Ca3(PO4)2 and 0.2M Mg(OH)2 was also found resistant to the clotting action of the antibacterial agent. The clotting action of the antibacterial agent was not affected by heating at 66 C for 60 min, nor by multiple passage through Seitz filters. The coagulation of citrated plasma proceeded most rapidly with 0.2M Tris(hydroxymethyl)-amino-methane buffer, pH 7 to 9, or distilled water as the antibacterial agent solvent; 0.2M phosphate buffer, pH 6 to 8, reduced the clotting action of the antibacterial agent, while 0.2M citrate-phosphate buffer, pH 4.0, or 0.2M carbonate-bicarbonate buffer, pH 10.0, inhibited entirely the clotting activity of this agent. 相似文献
7.
Luciano Galizia Gabriela I. Marino Alejandro Ojea Basilio A. Kotsias 《The Journal of membrane biology》2013,246(12):949-958
The regulation of the epithelial Na+ channel (ENaC) during cell swelling is relevant in cellular processes in which cell volume changes occur, i.e., migration, proliferation and cell absorption. Its sensitivity to hypotonically induced swelling was investigated in the Xenopus oocyte expression system with the injection of the three subunits of mouse ENaC. We used voltage-clamp techniques to study the amiloride-sensitive Na+ currents (INa(amil)) and video microscopic methodologies to assess oocyte volume changes. Under conditions of mild swelling (25 % reduced hypotonicity) inward current amplitude decreased rapidly over 1.5 min. In contrast, there was no change in current amplitude of H2O-injected oocytes to the osmotic insult. INa(amil) kinetics analysis revealed a decrease in the slower inactivation time constant during the hypotonic stimuli. Currents from ENaC-injected oocytes were not sensitive to external Cl? reduction. Neither short- nor long-term cytochalasin D treatment affected the observed response. Oocytes expressing a DEG mutant β-ENaC subunit (β-S518K) with an open probability of 1 had reduced INa(amil) hypotonic response compared to oocytes injected with wild-type ENaC subunits. Finally, during the hypotonic response ENaC-injected oocytes did not show a cell volume difference compared with water-injected oocytes. On this basis we suggest that hypotonicity-dependent ENaC inhibition is principally mediated through an effect on open probability of channels in the membrane. 相似文献
8.
Ce3+/Eu2+ co‐doped Na3Ca6(PO4)5 phosphors were prepared using a combustion‐assisted synthesis method. X‐Ray powder diffraction (XRD) analysis confirmed the formation of a Na3Ca6(PO4)5 crystal phase. Na3Ca6(PO4)5:Eu2+ phosphors have an efficient bluish‐green emission band that peaks at 489 nm, whereas Ce3+‐doped Na3Ca6(PO4)5 showed a bright emission band at 391 nm. Analysis of the experimental results suggests that enhancement of the Eu2+ emission intensity in co‐doped Na3Ca6(PO4)5:Eu2+,Ce3+ phosphors is due to a resonance‐type energy transfer from Ce3+ to Eu2+ ions, which is predominantly governed by an exchange interaction mechanism. These results indicate that Ce3+/Eu2+ co‐doped Na3Ca6(PO4)5 is potentially useful as a highly efficient, bluish‐green emitting, UV‐convertible phosphor for white‐light‐emitting diodes. Copyright © 2014 John Wiley & Sons, Ltd. 相似文献
9.
A. E. Ronco 《World journal of microbiology & biotechnology》1992,8(3):316-318
Lyophilized cells ofPhotobacterium phosphoreum, rehydrated in 2% (w/v) NaCl in 0.022M KH2PO4 at pH 7.0, were used for developing an assay to test the acute toxicity of organic and inorganic compounds. The standardized assay gave good reproducibility of results with 11 organic and four inorganic compounds. Results were compared with reported data obtained with other test organisms and are within their sensitivity ranges. Environmental screening of wastes from oil and petrochemical industries is discussed. 相似文献
10.
The NHE-1 isoform of the Na+/H+ exchanger is excessively activated in cardiac cells during ischemia. Hence NHE-1 specific inhibitors are being developed
since they could be of beneficial influence under conditions of cardiac ischemia and reperfusion. In this study, the Cytosensor™
microphysiometer was used to measure the potency of four new drug molecules, i.e., EMD 84021, EMD 94309, EMD 96785 and HOE
642 which are inhibitors of the isoform 1 of the Na+/H+ exchanger. The experiments were performed with Chinese hamster ovary cells (CHO K1) which are enriched in the NHE-1 isoform
of the Na+/H+ antiporter. The Na+/H+ exchanger was stimulated with NaCl and the rate of extracellular acidification was quantified with the Cytosensor. The proton
exchange rate was measured as a function of the NaCl concentration in the range of 10–138 mm NaCl stimulation. The proton exchange rate followed Michaelis-Menten kinetics with a K
M
= 30 ± 4 mm for Na+. Addition of either one of the four inhibitors decreased the acidification rate. The IC50 values of the four compounds could be determined as 23 ± 7 nm for EMD 84021, 5 ± 1 nm for EMD 94309, 9 ± 2 nm for EMD 96785 and 8 ± 2 nm for HOE 642 at 138 mm NaCl, in good agreement with more elaborate biological assays. The IC50 values increased with the NaCl concentration indicating competitive binding of the inhibitor. The microphysiometer approach
is a fast and simple method to measure the activity of the Na+/H+ antiporter and allows a quantitative kinetic analysis of the proton excretion rate.
Received: 3 September 1998/Revised: 20 November 1998 相似文献
11.
Kevin Strange 《The Journal of membrane biology》1989,107(3):249-261
Summary Ouabain had no effect on the volume of intercalated cells of DOCA-stimulated rabbit cortical collecting tubules, but caused principal cells to swell rapidly at an initial rate of 67% min., Principal cells swelled 133% then activated regulatory volume decrease mechanisms and shrank at an initial rate of –3%/min to a new volume 13% above control. The initial rate of ouabain swelling was completely inhibited by perfusate Na+ removal or reduced 95% by luminal addition of 10–5
m amiloride. Luminal peritubular, or bilateral Cl– removal each caused cell shrinkages of 10% and reduced the rate of ouabain swelling by 70, 85, and 99%, respectively. The presence of an apical Cl– transport step in principal cells was confirmed by increasing luminal K+ from 5 to 53mm, which caused cell swelling of 22%. This volume increase was completely blocked by luminal Cl– removal, but was unaffected by peritubular Cl– substitution. Perfusion of CCT with 0.1mm acetazolomide, 0.1mm DPC or 0.5mm SITS caused principal cell shrinkages of 7–9% and reduced the rate of ouabain swelling by 60, 70, and 40%, respectively. The initial rate of ouabain swelling was inhibited 70% by bilateral CO2/HCO3 removal and 50% by whole animal acid loading. Taken together these results demounstrate that ouabain swelling is due to cellular NaCl accumulation and that Na+ enters the cell primarily through apical Na+ channels. Cellular Cl– entry occurs at least partially through the apical membrane and may be mediated by a Cl–/HCO
3
–
exchanger. Brief (45–90 sec) exposure of principal cells to ouabain is associated with a rapid inhibition of Na+ and/or Cl– entry steps, whereas long-term (>5min) ouabvain exposure completely blocks one or both of these transport pathways. 相似文献
12.
Summary A fraction of erythrocyte Band 3 (M
r
, 93,000) glycoprotein that demonstrates decreased autophosphorylation in membranes from myotonic muscular dystrophy patients is demonstrated. Sequential affinity chromatography of Triton X-100 solubilized erythrocyte membrane proteins separated three specifically retained glycoprotein fractions on a Ricin Communis I-Sepharose 4B column. One fraction contains a portion of the major sialoglycoprotein (apparentM
r
, 78,000) and is specifically eluted from the column by 10mm NaCl and 100mm d-galactose (10/100). The two other glycoprotein fractions are eluted by 100mm NaCl, 10mm d-galactose (100/10) and 100mm NaCl, 100mm d-galactose (100/100). The composition of both fractions contains greater than 95% Band 3 (apparentM
r
, 93,000) glycoprotein.The quantities of glycoprotein in each fraction obtained from erythrocytes of myotonic dystrophy patients did not differ from the quantities obtained from control erythrocytes. Following endogenous protein kinase incubations of ghosts with [-32P]ATP, the specific [32P] phosphorylation of the 10/100 and 100/10 fractions are identical. The 100/100 fraction, which makes up approximately 3% of the total erythrocyte membrane protein, demonstrates a different pattern for myotonic dystrophy patients; specific phosphorylation was reduced by 50% relative to activity in control experiments. These findings are consistent with previous experiments that demonstrated decreased autophosphorylation of the glycoprotein portion of Band 3 (Roses & Appel, 1975,J. Membrane Biol.
20: 51) and are consistent with the autosomal dominant mode of inheritance in this disease. 相似文献
13.
We studied the relationship between changes in intracellular pH (pH
i
), intracellular Ca2+([Ca2+]
i
) and charybdotoxin sensitive (CTX) maxi-K+ channels occurring after modest `physiological' swelling in guinea pig jejunal villus enterocytes. Villus cell volume was
assessed by electronic cell sizing, and pH
i
and [Ca2+]
i
by fluorescence spectroscopy with 2,7, biscarboxyethyl-5-6-carboxyfluorescein and Indo-1, respectively. In a slightly (0.93
× isotonic) hypotonic medium, villus cells swelled to the same size they would reach during d-glucose or l-alanine absorption; the subsequent Regulatory Volume Decrease (RVD) was prevented by CTX. After the large volume increase
in a more hypotonic (0.80 × isotonic) medium, RVD was unaffected by CTX. After modest swelling associated with 0.93 × isotonic
dilution, the pH
i
alkalinized but N-5-methyl-isobutyl amiloride (MIA) prevented this ΔpH
i
and the subsequent RVD. Even in the presence of MIA, alkalinization with added NH4Cl permitted complete RVD which could be inhibited by CTX. The rate of 86Rb efflux which also increased after this 0.93 × isotonic dilution was inhibited an equivalent amount by CTX, MIA or Na+-free medium. Modest swelling transiently increased [Ca2+]
i
and Ca2+-free medium or blocking alkalinization by MIA or Na+-free medium diminished this transient increase an equivalent amount. RVD after modest swelling was prevented in Ca2+-free medium but alkalinization still occurred. After large volume increases, alkalinization of cells increased [Ca2+]
i
and volume changes became sensitive to CTX. We conclude that both alkalinization of pH
i
and increased [Ca2+]
i
observed with `physiological' volume increase are essential for the activation of CTX-sensitive maxi-K+ channels required for RVD.
Received: 30 March 1999/Revised: 6 July 1999 相似文献
14.
M. Iino S. Ciani K. Tsuzuki S. Ozawa Y. Kidokoro 《The Journal of membrane biology》1997,155(2):143-156
Ion permeation properties of the mouse e2/ζ1 NMDA receptor channel expressed in Xenopus oocytes were studied using the outside-out patch-clamp technique. In symmetrical Na+ solutions, the single-channel I-V relations were almost linear at low electrolyte concentrations, but rectified inwardly for Na+ concentrations above 50 mm. In symmetrical Na+ solutions, the ``zero-current conductance' increased with Na+ concentration and saturated according to a hyperbolic curve, the half-maximal saturating activity, K
M
(Na), being 14.2 mm and the maximal conductance, G
max(Na), 53.9 pS. When Ca2+ was present with Na+ in the external solution, the single-channel current was lower than in pure Na+, although the reversal potential indicated a higher permeability for Ca2+ than for Na+. Using ion activities, PCa/PNa was found to be about 17. The I-V data were fitted with a model based on the Eyring's rate theory, assuming a one-ion pore with three energy barriers and two
sites. The K
M
(Ca) and G
max (Ca) were 76.5 μm and 21.2 pS, respectively. According to the estimated rate constants, K
M
for Ca2+ is mainly determined by the binding strength of a site located 80% away from the channel opening at the external membrane-solution
interface, a position similar to that postulated previously for the Mg2+ blocking site.
Received: 3 May 1996/Revised: 25 September 1996 相似文献
15.
M. Aschner K. J. Mullaney M. N. Fehm D. E. Wagoner Jr. D. Vitarella 《Cellular and molecular neurobiology》1994,14(6):637-652
Summary 1. MC has been shown to inhibit the uptake ofl-glutamate and increased-aspartate release from preloaded astrocytes in a dose-dependent fashion.2. Two sulfhydryl (SH-)-protecting agents; reduced glutathione (GSH), a cell membrane-nonpenetrating compound, and the membrane permeable dithiothreitol (DTT), have been shown consistently to reverse the above effects. MC-inducedd-aspartate release is completely inhibited by the addition of 1 mM DTT or GSH during the actual 5-min perfusion period with MC (5µM); when added after MC treatment, DTT fully inhibits the MC-inducedd-aspartate release, while GSH does not.3. Neither DTT nor GSH, in the absence of MC, have any effect on the rate of astrocyticd-aspartate release. Other studies demonstrate that although MC treatment (5µM) does not induce astrocytic swelling, its addition to astrocytes swollen by exposure to hypotonic medium leads to their failure to volume regulate.4. Omission of calcium from the medium greatly potentiates the effect of MC on astrocyticd-aspartate release, an effect which can be reversed by cotreatment of astrocytes with the dihydropyridine Ca2+-channel antagonist nimodipine (10µM), indicating that one possible route of MC entry into the cells is through voltage-gated L-type channels. 相似文献
16.
A dual-wavelength fluorimeter was constructed, which used two light emitting diodes (LEDs) to excite the fluorescence dye RH 421 alternately with two different wavelengths. The ratio of the emissions at the two excitation wavelengths provided a drift-insensitive signal, which allowed detection of very small changes of the fluorescence intensity. Those small changes were induced by ion binding and release in conformation E1 of the Na,K-ATPase. Titration experiments were performed to determine equilibrium dissociation constants (± standard deviation) for each step in the complete binding and release sequence: 0.12 ± 0.01 mM (E2(K2) KE1), 0.08 ± 0.01 mM (KE1 E1), 3.0 ± 0.2 mM (NaE1 E1), 5.2 ± 0.4 mM (Na2E1 NaE1) and 6.5 ± 0.4 mM (Na3E1 Na2E1) at pH 7.2 and T=16°C. These numbers show that the affinities of the binding sites exposed to the cytoplasm, are higher for K+ than for Na+ ions, similar to what was found on the extracellular side. The physiological requirement for extrusion of Na+ from the cytoplasm, and for import of K+ from the extracellular medium seems to be facilitated not by favorable binding affinities in state E1 but by the two ATP-driven reaction steps of the cycle, E2(K2) + ATP K2E1 · ATP and Na3E1 · ATP (Na3) El-P, which border the ion exchange reactions at the binding sites in conformation E1.
Correspondence to: H.-J. Apell 相似文献
17.
Summary A vanadate-sensitive H+-translocating ATPase isolated from red beet plasma membrane has been solubilized in active form and successfully reconstituted into artificial proteoliposomes. The H+-ATPase was solubilized in active form with deoxycholate, CHAPSO or octylglucoside in the presence of glycerol. Following detergent removal by gel filtration and reconstitution into proteoliposomes, ATP:Mg-dependent H+ transport could be measured as ionophore-reversible quenching of acridine orange fluorescence. Solubilization resulted in a three-to fourfold purification of the plasma membrane ATPase, with some additional enrichment of specific activity following reconstitution. H+ transport activity was inhibited half-maximally between 1 and 5 M vanadate (Na3VO4) and nearly abolished by 100 M vanadate. ATPase activity of native plasma membrane showed aK
i
for vanadate inhibition of 9.5 M, and was inhibited up to 80% by 15 to 20 M vanadate (Na3VO4). ATPase activity of the reconstituted vesicles showed aK
i
of 2.6 M for vanadate inhibition. The strong inhibition by low concentrations of vanadate indicates a plasma membrane rather than a mitochondrial or tonoplast origin for the reconstituted enzyme. 相似文献
18.
Tomonori Takashina Kiyotaka Otozati Tetsuo Hamamoto Koki Horikoshi 《Biodiversity and Conservation》1994,3(7):632-642
Halophilic and halotolerant bacteria were isolated from soil samples of a Japanese salt field, an environment where salt concentrations vary annually. From 1 g of each of the five samples collected, over 1×103 bacterial colonies (colony forming units (cfu)g-1) grew on agar medium containing 2M Na+. In contrast, 0–4 bacterial colonies (cfu g-1) were observed on agar medium containing 4M Na+. Two of the five samples contained numerous bacteria (102–103 cfu g-1) capable of growth on a 2M Na+ alkaline (pH=9.5) medium, while few bacterial colonies were observed from the other three samples. Only one of the five samples was shown to contain bacteria capable of growth on a 4M Na+ alkaline medium. Most of the bacteria isolated on 4M Na+ agar were eubacteria, but one extreme halophile (TR-1, already described as Haloarcula japonica JCM7785) was also isolated. The 16S rRNA sequence of TR-1 was determined and shows high homology (94.4–98.5%) to Ha. marismortui and Ha. sinaiiensis. These results suggested that: 1) environments with seasonally varying salinity can harbour halotolerants as well as halophiles and, 2) closely related halophiles can be isolated from geographically distant habitats. 相似文献
19.
Anthony F. Almeida Simon M. Jarvis James D. Young Alan R.P. Paterson 《FEBS letters》1984,176(2):444-448
Nitrobenzylthioniosine (NBMPR), a potent and specific inhibitor of nucleoside transport, is bound reversibly by high affinity sites on nucleoside transporter proteins of erythrocyte membranes and, upon photoactivation, NBMPR molecules become covalently bonded to the sites. This study showed that [3H]NBMPR molecules reversibly bound to intact S49 and L5178Y mouse lymphoma cells became covalently bound upon exposure to UV light. Electrophoretic analysis of plasma membrane fractions from the labelled cells showed that 3H was present in polypeptides which migrated as a major band with an apparent Mr of 45000–65000. 相似文献
20.
Histochemical demonstration of carbonic anhydrase activity 总被引:14,自引:4,他引:10
Holger P. J. Hansson 《Histochemistry and cell biology》1967,11(2):112-128
Summary Freeze-dried frozen sections are floated on the surface of the freshly prepared incubation mixture (CoSO4 1.75 × 10–3
M, H2SO4 5.3 × 10–2
M, NaHCO3 1.57 × 10–2
M and KH2PO4 1.17 to 11.7 × 10–3
M; demonstration of weak activity requires high phosphate). A compound containing cobalt and phosphorous precipitates at carbonic anhydrase sites and is converted to CoS. Adequate staining requires only 2–10 minutes of incubation. Actazolamide inhibits the staining reaction in specific concentrations. Actazolamidein vivo, 20 mg/kgi.v. to mice 30 minutes before sacrifice also inhibited the staining. The proportion phosphorous in the specific precipitate increases with KH2PO4 of the medium (shown by the addition of60Co and32P). An explanation of the reaction mechanism is given, based on the catalyzed loss of CO2 in the surface layer. The inclusion of phosphate in the medium makes this modification ofHäusler's method so sensitive that it shows carbonic anhydrase activity in for instance stratum spinosum of the skin.This investigation was supported by grants from the Medical Faculty, University of Uppsala and from the U.S. National Institutes of Health (Grant NB 3060 to E.Bárány). 相似文献