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1.
Fe2+ transport in plants has been difficult to quantify because of the inability to control Fe2+ activity in aerated solutions and non-specific binding of Fe to cell walls. In this study, a Fe(II)-3-(2-pyridyl)-5,6-diphenyl-1,2,4-triazine-4[prime]4"-disulfonic acid buffer system was used to control free Fe2+ in uptake solutions. Additionally, desorption methodologies were developed to adequately remove nonspecifically bound Fe from the root apoplasm. This enabled us to quantify unidirectional Fe2+ influx via radiotracer (59Fe) uptake in roots of pea (Pisum sativum cv Sparkle) and its single gene mutant brz, an Fe hyperaccumulator. Fe influx into roots was dramatically inhibited by low temperature, indicating that the measured Fe accumulation in these roots was due to true influx across the plasma membrane rather than nonspecific binding to the root apoplasm. Both Fe2+ influx and Fe translocation to the shoots were stimulated by Fe deficiency in Sparkle. Additionally, brz, a mutant that constitutively exhibits high ferric reductase activity, exhibited higher Fe2+ influx rates than +Fe-grown Sparkle. These results suggest that either Fe deficiency triggers the induction of the Fe2+ transporter or that the enhanced ferric reductase activity somehow stimulates the activity of the existing Fe2+ transport protein.  相似文献   

2.
In vivo kinetics of mucosal uptake of luminal 59Fe2+ by tied segments of normal mouse duodenum are characterised by a Km of approx. 100 μM and a Vmax of approx. 9 pmol/min per mg wet weight of intestine. These values were determined at pH 7.25 in the presence of excess sodium ascorbate. Studies with luminal Fe2+ concentrations of 100 μM reveal: (1) uptake is relatively independent of ascorbate: Fe ratio and luminal pH and (2) uptake is potently inhibited by 1 mM Co2+ or Mn2+ and large luminal NaCl concentrations but not by Ca2+. 3 days of hypoxia (0.5 atmospheres) yields no significant increase in subsequent total mucosal uptake by in vivo tied segments while uptake is significantly reduced by semi-starvation. Quantitative comparison of in vivo mucosal uptake with subsequent determination of isolated brush-border membrane 59Fe2+ transport in individual mice reveals a positive correlation (P < 0.01) between the two parameters. These results, in conjunction with studies of isolated mouse duodenal brush-border membrane (Simpson, R.J. and Peters, T.J. (1985) Biochim. Biophys. Acta, 814, 381–388 and (1986) Biochim. Biophys. Acta 856, 109–114) suggest that the Fe2+ transport properties of isolated brush-border membrane are quantitatively adequate to explain in vivo mucosal uptake in normal and hypoxic mice at Fe2+ concentrations up to 100 μM.  相似文献   

3.
Initial rates of 59Fe3+ uptake by mouse duodenal fragments (in vitro) and tied-off duodenal segments (in vivo) have been characterised for control and hypoxic animals. 59Fe3+ uptake by duodenal fragments was rapid, selective and dependent on medium Fe3+-nitrilotriacetate concentration. Most of the 59Fe3+ uptake (70-75%) occurred via the mucosal route and was dependent on the metabolic state of the tissue. Mucosal uptake showed an adaptive increase following exposure of animals to 3 days hypoxia; the enhancement was due to a 2-3-fold increase in Vmax app, without any significant changes in the Km app. Studies of upper small intestine transit times showed a mean residence time of 4-5 min for 59Fe-labelled mouse chow, emphasising the importance of initial uptake measurements. Time courses for in vivo total mucosal uptake exhibited linearity over a wide variety of absorption rates after correction for the permeation by intact metal-chelate complex. The corrected uptake showed a hyperbolic dependence on medium Fe3+-nitrilotriacetate concentration. Kinetic studies revealed a 2-3-fold increase in total mucosal uptake in hypoxia. Mucosa-to-carcass transfer of 59Fe was also markedly increased by chronic hypoxia. The in vitro system exhibits similar qualitative and quantitative kinetics for Fe3+ transport via the mucosal membrane to those obtained in vivo. The results observed in vitro are thus valid and provide a convenient method for further studies on Fe3+ transport in animals and in man.  相似文献   

4.
A study has been made on the effect of primary leaves on iron (Fe) distribution in the shoot. Bean (Phaseolus vulgaris L.) seedlings were precultured in nutrient solution with 8×10-5 M FeEDTA for 4 days, and then grown further with either 8×10-5 M FeEDTA (+Fe) or without Fe supply (-Fe) for another 5 days. Thereafter, both +Fe and -Fe plants were treated in three different ways: undisturbed; one primary leaf removed; or one primary leaf shaded, starting two hours before supply 59FeEDTA to the roots. The +Fe plants were supplied with 8×10-5 M 59FeEDTA, and the -Fe plants with only 1×10-6 M 59FeEDTA. After 1 to 8 hour uptake periods, plants were harvested and 59Fe in different organs was determined. Removal or shading of one primary leaf did not affect 59Fe uptake by roots and 59Fe translocation to the shoot in +Fe plants. In the -Fe plants, however, removal of one primary leaf decreased 59Fe uptake by roots, whereas shading of one primary leaf had no effect on 59Fe uptake but slightly enhanced 59Fe translocation from roots to the shoot. The quantity of 59Fe in primary leaves was positively correlated with quantity of 59Fe in the stem in the -Fepplants, but not in the +Fe plants. In both, the +Fe and -Fe plants, the quantity of 59Fe in the shoot apex was positively correlated with 59Fe in primary leaves. The results suggest that irrespective of the Fe nutritional status of plants, the source of Fe for the shoot apex is Fe retranslocated from primary leaves.  相似文献   

5.
6.
In the management of lake eutrophication, the regulation effect of Fe is considered, in addition to the controlling nitrogen- and phosphorus input. Based on the “Fe hypothesis”, this paper tentatively ap-plied plant spectral response to the remote sensing early-warning mechanism of lake eutrophication. A laboratory water culture experiment with rice (Oryza sativa L.) was conducted to study Fe uptake by plants and the chlorophyll concentration and visible-near infrared spectrum of vegetable leaves as well as their interrelations under Fe2+ stress. Three spectral indices, i.e., A1 (integral value of the changes of spectral reflectivity in the range 460―670 nm under Fe2+ stress), A2 (integral value of the changes of spectral reflectivity in the range of 760―1000 nm under Fe2+ stress) and S (blue-shift range of red edge curve under Fe2+ stress), were used to establish quantitative models about the relationships between the rice leaf spectrum and Fe2+ stress. With the increase of Fe2+ in a culture solution, the Fe content in rice plants increased, while the chlorophyll concentration in vegetative leaves decreased. The spectral reflectivity of vegetable leaves increased in the visible light band but decreased in the near infrared band, and the blue-shift range of the red edge curve increased. The indices A1, A2 and S all had sig-nificant correlations with the Fe content in rice leaves, the correlation coefficient being respectively 0.951 (P < 0.01), −0.988 (P < 0.01) and 0.851 (P < 0.01), and simulated (multiple correlation coefficients R2 > 0.96) and predict the Fe level in rice leaves.  相似文献   

7.
Summary Experiments were performed to obtain information on: (i) the specific properties of Ca2+ binding and transport in yeast (ii) the relationship between both parameters; (iii) similarities to or differences from other biological systems as measured by the effects of inhibitors; and (iv) the effects of mono and divalent cations, in order to get some insight on the specificity and some characteristics of the mechanism of the transport system for divalent cations in yeast.The results obtained gave some kinetic parameters for a high affinity system involved in the transport of Ca2+ in yeast. These were obtained mainly by considering actual concentrations of Ca2+ in the medium after substracting the amounts bound to the cell. Ak m of 1.9 m and aV max of 1.2 nmol (100 mg·3 min)–1 were calculated.The effects of some inhibitors and other cations on Ca2+ uptake allow one to postulate some independence between binding and transport for this divalent cation.Of the inhibitors tested, only lanthanum seems to be a potent inhibitor of Ca2+ uptake in yeast.The effects of Mg2+ on the uptake of Ca2+ agree with the existence of a single transport system for both divalent cations.The actions of Na+ and K+ on the transport of Ca2+ offer interesting possibilities to study further some of the mechanistic properties of this transport system for divalent cations.  相似文献   

8.
Reduction and transport of Fe from siderophores   总被引:1,自引:0,他引:1  
Soils contain siderophores produced by bacteria and fungi; however, the role of siderophores in Fe nutrition of plants is uncertain. The Strategy I plant cucumber (Cucumis sativus L.) was used in an investigation of ferric chelate reduction activity and uptake and transport of Fe from ferric hydroxyethylethylenetriacetic acid (FeHEDTA) and ferric N,N–di–(2–hydroxybenzoyl)–ethylenediamine– N,N-diacetic acid (FeHBED) and the hydroxamate siderophores, ferric rhodotorulic acid (FeRA) and ferric ferrioxime B (FeFOB). Cucumber seedlings were grown in a hydroponic medium without Fe or supplied with 10 M FeHEDTA. Iron-deficient cucumber roots readily reduced FeHEDTA, while Fe-sufficient roots had low levels of ferric chelate reduction activity. The siderophore FeRA was reduced by Fe-deficient roots at 8% of the rate of FeHEDTA, while FeFOB was not reduced. The highly stable synthetic chelate FeHBED was reduced at 16% the rate of FeHEDTA. Fe transport to shoots by Fe-deficient seedlings from the slowly reducible complexes 59FeRA and 59FeHBED was, respectively, 74% and 73% of that transported from 59FeHEDTA. The ferrous complexing agent, bathophenanthrolinedisulfonic acid (BPDS), had a strong inhibitory effect on uptake and transport of Fe from 59FeHEDTA or 59FeRA into shoots. An average of 11% as much Fe was transported to shoots of Fe-deficient seedlings from 59FeFOB as from 59FeHEDTA. Neither the Fe nutritional status of the seedlings nor the presence of BPDS influenced the uptake and transport of Fe from 59FeFOB. It is concluded that cucumber roots may take up substantial amounts of Fe from FeRA and FeHBED following reduction, while small amounts of Fe may be taken up from FeFOB by a mechanism not involving reduction of the ferric siderophore at the root surface.  相似文献   

9.
Abstract

In order to examine whether chiral metal complexes can be used to discriminate between right- and left-handed DNA conformational states we have studied the enantioselective interactions of Fe(phen)3 2+ and Ru(phen)3 2+ (phen = 1,10-phenanthroline)with poly(dGm5dC) under B- and Z-form conditions. With the inversion-labile Fe(phen)3 2+, enantioselectivity leads to shifts in the diastereomeric binding equilibria. This effect, known as the “Pfeiffer effect” (1–4), is monitored as a slowly emerging circular dichroism of the solution, corresponding to a net excess of the favoured enantiomer. With Ru(phen)3 2+, which is stable to intramolecular inversion, the difference in DNA-binding strengths of the enantiomers results in an excess of the less favoured enantiomer in the bulk solution. This excess is detected in the dialysate of the DNA/metal complex solution. With both complexes we find that the Δ-enantiomer is favoured when the polynucleotide adopts the B-form, as previously shown, but also when it initially adopts the Z-form conformational state.

This observation, together with evidence from UV-circular dichroism and binding data, indicates that the binding of these metal complexes induces a Z- to B-form transition in Z- form poly(dGm5dC). Consequently, neither of the studied chiral DNA-binders can easily be used to discriminate the DNA handedness.  相似文献   

10.
Cobalt nanoparticles (CoNPs) released from hip joint implants are known to have a toxic effect on several organs probably through increasing reactive oxygen species (ROS). Ferrous ion (Fe2+) is well-known to enhance oxidative stress by catalysing the production of ROS. However, in our pilot study, we found that Fe2+ conversely inhibited the ROS production induced by CoNPs. To elucidate the underlying mechanism, the present study treated vascular endothelial HUVEC and HMEC-1 cells with CoNPs alone or in combination with ferrous lactate [Fe(CH3CHOHCOO)2], ferrous succinate [Fe(CH2COO)2], and ferrous chloride (FeCl2). CoNP toxicity was evaluated by measuring cell viability, rate of apoptosis and lactose dehydrogenase (LDH) release, and intracellular ROS levels. Treatment with CoNPs decreased cell viability, LDH release, and ROS production and increased apoptosis. CoNPs increased hypoxia-inducible factor-1α (HIF-1α) protein level and mRNA levels of vascular endothelial growth factor (VEGF) and glucose transporter 1 (GLUT1) downstream of HIF-1α signalling. Silencing HIF-1α attenuated CoNP toxicity, as seen by recovery of cell viability, LDH release, and ROS levels and reduced apoptosis. CoNPs caused a pronounced reduction of Fe2+ in cells, but supplementation with Fe(CH3CHOHCOO)2, Fe(CH2COO)2, and FeCl2 restored Fe2+ levels and inhibited HIF-1α activation. Moreover, all three Fe2+-containing agents conferred protection from CoNPs; Fe(CH3CHOHCOO)2 and Fe(CH2COO)2 more effectively than FeCl2. In summary, the present study revealed that CoNPs exert their toxicity on human vascular endothelial cells by depleting intracellular Fe2+ level, which causes activation of HIF-1α signalling. Supplements of Fe2+, especially in the form of Fe(CH3CHOHCOO)2 and Fe(CH2COO)2, mitigated CoNP toxicity.  相似文献   

11.
The metal ions Zn2+, Cu2+, and Fe2+ play a significant role in the aggregation mechanism of Aβ peptides. However, the nature of binding between metal and peptide has remained elusive; the detailed information on this from the experimental study is very difficult. Density functional theory (dft) (M06‐2X/6‐311++G (2df,2pd) +LANL2DZ) has employed to determine the force field resulting due to metal and histidine interaction. We performed 200 ns molecular dynamics (MD) simulation on Aβ1‐42‐Zn2+, Aβ1‐42‐Cu2+, and Aβ1‐42‐Fe2+ systems in explicit water with different combination of coordinating residues including the three Histidine residues in the N‐terminal. The present investigation, the Aβ1‐42‐Zn2+ system possess three turn conformations separated by coil structure. Zn2+ binding caused the loss of the helical structure of N‐terminal residues which transformed into the S‐shaped conformation. Zn2+ has reduced the coil and increases the turn content of the peptide compared with experimental study. On the other hand, the Cu2+ binds with peptide, β sheet formation is observed at the N‐terminal residues of the peptide. Fe2+ binding is to promote the formation of Glu22‐Lys28 salt‐bridge which stabilized the turn conformation in the Phe19‐Gly25 residues, subsequently β sheets were observed at His13‐Lys18 and Gly29‐Gly37 residues. The turn conformation facilitates the β sheets are arranged in parallel by enhancing the hydrophobic contact between Gly25 and Met35, Lys16 and Met35, Leu17 and Leu34, Val18 and Leu34 residues. The Fe2+ binding reduced the helix structure and increases the β sheet content in the peptide, which suggested, Fe2+ promotes the oligomerization by enhancing the peptide‐peptide interaction. Proteins 2016; 84:1257–1274. © 2016 Wiley Periodicals, Inc.  相似文献   

12.
K+-dependent Na+-Ca2+ exchangers (NCKXs) play an important role in Ca2+ homeostasis in many tissues. NCKX proteins are bi-directional plasma membrane Ca2+-transporters which utilize the inward Na+ and outward K+ gradients to move Ca2+ ions into and out of the cytosol (4Na+:1Ca2+ + 1 K+). In this study, we carried out scanning mutagenesis of all the residues of the highly conserved α-1 and α-2 repeats of NCKX2 to identify residues important for K+ transport. These structural elements are thought to be critical for cation transport. Using fluorescent intracellular Ca2+-indicating dyes, we measured the K+ dependence of transport carried out by wildtype or mutant NCKX2 proteins expressed in HEK293 cells and analyzed shifts in the apparent binding affinity (Km) of mutant proteins in comparison with the wildtype exchanger. Of the 93 residue substitutions tested, 34 were found to show a significant shift in the external K+ ion dependence of which 16 showed an increased affinity to K+ ions and 18 showed a decreased affinity and hence are believed to be important for K+ ion binding and transport. We also identified 8 residue substitutions that resulted in a partial loss of K+ dependence. Our biochemical data provide strong support for the cation binding sites identified in a homology model of NCKX2 based on crystal structures reported for distantly related archaeal Na+-Ca2+ exchanger NCX_Mj. In addition, we compare our results here with our previous studies that report on residues important for Ca2+ and Na+ binding. Supported by CIHR MOP-81327.  相似文献   

13.
The addition of nanomolar concentrations of free Fe2+, Mn2+, or Co2+ to rat liver plasma membranes resulted in an activation of ATP hydrolysis by these membranes which was not additive with the Ca2+-stimulated ATPase activity coupled to the Ca2+ pump. Detailed analysis showed that, if fact, (i) as for the stimulation of (Ca2+-Mg2+)-ATPase by Ca2+, activation of ATP hydrolysis by Fe2+, Mn3+, or Co2+ followed a cooperative mechanism involving two ions; (ii) two interacting sites for ATP were involved in the activation of both Fe2+- and Ca2+-stimulated ATPase activities; (iii) micromolar concentrations of magnesium caused the same dramatic inhibition of both activities; and (iv) the subcellular distribution of Fe2+-activated ATP hydrolysis activity corresponded to that of plasma membrane markers. This suggests that the (Ca2+-Mg2+)-ATPase might be stimulated not only by Ca2+, but also by Fe2+, Mn2+, or Co2+. However, interaction of (Ca2+-Mg2+)-ATPase with Fe2+, Mn2+, or Co2+ inhibited the Ca2+ pump activity. Furthermore, neither the formation of the phosphorylated intermediate of (Ca2+-Mg2+)-ATPase, nor ATP-dependent (59Fe) uptake could be detected in the presence of Fe2+ concentrations which stimulated ATP hydrolysis. We conclude that: (i) under the influence of certain metal ions, the Ca2+ pump in the liver plasma membrane may be switched to an uncoupled state which displays ATP hydrolysis activity, but does not insure ion transport; (ii) therefore the Ca2+ pump in liver plasma membranes specifically insures Ca2+ transport.  相似文献   

14.
Abstract

Clotrimazole is an antimycotic imidazole derivative that interferes with cellular Ca2+ homeostasis. This study examined the effect of clotrimazole on cytosolic Ca2+ concentrations ([Ca2+]i) and viability in HA59T human hepatoma cells. The Ca2+-sensitive fluorescent dye fura-2 was applied to measure [Ca2+]i. Clotrimazole induced [Ca2+]i rises in a concentration-dependent manner. The response was reduced by removing extracellular Ca2+. Clotrimazole-evoked Ca2+ entry was suppressed by store-operated channel inhibitors (nifedipine, econazole and SK&F96365) and protein kinase C modulators (GF109203X and phorbol, 12-myristate, 13-acetate). In Ca2+-free medium, incubation with the endoplasmic reticulum Ca2+ pump inhibitor 2,5-di-tert-butylhydroquinone abolished clotrimazole-induced [Ca2+]i rise. Inhibition of phospholipase C with U73122 abolished clotrimazole-induced [Ca2+]i rise. At 10–40?µM, clotrimazole inhibited cell viability, which was not reversed by chelating cytosolic Ca2+. Clotrimazole at 10 and 30?µM also induced apoptosis. Collectively, in HA59T cells, clotrimazole-induced [Ca2+]i rises by evoking phospholipase C-dependent Ca2+ release from the endoplasmic reticulum and Ca2+ entry via store-operated Ca2+ channels. Clotrimazole also caused apoptosis.  相似文献   

15.
The effects of ascorbic acid, sodium citrate, and sodium bicarbonate on59Fe-transferrin,54Mn-transferrin, and65Zn-transferrin uptake by the receptors disposed of plasma membrane isolated from lactating mouse mammary gland cells have been investigated. The effect of 10-2 mol/L ascorbic acid alone and in combination with NaHCO3 on the59Fe-transferrin uptake is significant and positive.54Mn-transferrin and65Zn-transferrin binding to the cell receptors are influenced optimally by 0.5 mol/L sodium bicarbonate. Sodium citrate alone or in combination with other substances always has a negative effect on binding of these three metals. It is suggested that a precise mechanism may exist with large possibilities to rearrange metal uptake and its transport from blood to milk.  相似文献   

16.
The inositol 1,4,5-trisphosphate receptor/channel (IP3R) is a major regulator of intracellular Ca2+ signaling, and liberates Ca2+ ions from the endoplasmic reticulum in response to binding at cytosolic sites for both IP3 and Ca2+. Although the steady-state gating properties of the IP3R have been extensively studied and modeled under conditions of fixed [IP3] and [Ca2+], little is known about how Ca2+ flux through a channel may modulate the gating of that same channel by feedback onto activating and inhibitory Ca2+ binding sites. We thus simulated the dynamics of Ca2+ self-feedback on monomeric and tetrameric IP3R models. A major conclusion is that self-activation depends crucially on stationary cytosolic Ca2+ buffers that slow the collapse of the local [Ca2+] microdomain after closure. This promotes burst-like reopenings by the rebinding of Ca2+ to the activating site; whereas inhibitory actions are substantially independent of stationary buffers but are strongly dependent on the location of the inhibitory Ca2+ binding site on the IP3R in relation to the channel pore.  相似文献   

17.
Zn2+ inhibits the binding of [59Fe]lactoferrin to neutrophilic leucocytes. The inhibiting effect is proportional to zinc concentration in the range 10-330 mumol/l. Zn2+ inhibits the [59Fe]lactoferrin binding to the colostral cells in the same degree as PMN. The inhibiting effect of Zn2+ on [59Fe]lactoferrin binding to neutrophilic leucocytes is equal to those of non-labelled lactoferrin and transferrin. Fe2+ and Cu2+ does not have such effect on binding of [59Fe]lactoferrin to the PMN leucocytes.  相似文献   

18.
Iron availability to plants is often limited when soil pH is 7 or higher. In C rich, but Fe limiting environments, microorganisms may produce organic chelators that complex Fe and increase its availability to plants. Seedlings of soybean (Glycine max L.) and oat (Avena sativa L.) plants, with Fe-efficient or inefficient uptake mechanisms, were grown in an Fe free nutrient solution at pH 7.5. Experiments (using a complete factorial design) were conducted in which these seedlings were transferred to a fresh nutrient solution and treated with Fe sources (FeCl3, FeEDDHA, and Fe complexed with chelators produced by compost microorganisms (CCMs) after their enrichment in an Fe free, glucose medium), Fe concentrations (0 and 6.7 M) and antibiotic (0 and 100 mg streptomycin L-1). Dry weight of soybean plants and Fe uptake were significantly (P 0.05) higher when Fe was supplied as 59FeCCM than as59 FeCl3 and similar to when Fe was supplied as59 FeEDDHA. Dry weight of the Fe-inefficient Tam 0-312 oat cultivar was also significantly higher when Fe was supplied as FeCCM. Fe uptake by oat, when supplied as 59FeCCM, was twice that for59 FeEDDHA and 59FeCl3. Chlorophyll concentration in plants supplied with FeCCM and FeEDDHA was significantly greater (P 0.05) than in minus Fe control plants and in FeCl3 supplied plants. Activities of catalase and peroxidase, measured as indicators of Fe nutrition in soybean and oats, were generally increased when Fe was supplied with FeCCM as compared to the other Fe sources. The experimental conditions in which the CCMs were produced are similar to those in soil after amendment with manures or other readily available organic materials. These CCMs can bind with Fe, even under slightly alkaline conditions, and effectively improve Fe nutrition of soybean and oat.  相似文献   

19.
Biochemical and kinetic properties under identical substrate and reaction conditions were obtained for an ATP-dependent Ca2+ pump and (Ca2+ + Mg2+)-ATPase in synaptosome membrane vesicles prepared from the brain of the moth, Mamestra configurata. Both the ATP-dependent Ca2+ pump and (Ca2+ + Mg2+)-ATPase had single, high-affinity binding sites for ATP (Km = 14 and 116 μM, respectively), Ca2+free (Km = 0.13 nM and 0.072 nM, respectively), and Mg2+ (Km = 1.1 mM and 0.07 mM, respectively). Both systems were relatively little affected by K+ and were insensitive to ouabain, an inhibitor of (Na+ + K+)-ATPase. The results indicate that the ATP-dependent Ca2+ pump and (Ca2+ + Mg2+)-ATPase are functionally coupled in synaptic membranes and constitute a mechanism for Ca2+ transport in the brain of M. configurata. Although moth brain (Ca2+ + Mg2+)-ATPase is maximally active at nanomolar concentrations of free calcium ion, the enzyme retains at least one-half of its maximal activity at micromolar calcium concentrations, indicating either that the enzyme has two binding sites for calcium (a high-affinity site at nanomolar Ca2+free and a low-affinity site at micromolar Ca2+free), or that there are two enzymes with high and low affinity for calcium, respectively. Calcium extrusion from brain neurones of M. configurata may operate in a two-stage, concentration-dependent process in which a first stage, low-affinity pump reduces intraneuronal calcium to a concentration at which a second stage, high-affinity pump becomes activated.  相似文献   

20.

Fe(II) cations bind with high efficiency and specificity at the high-affinity (HA), Mn-binding site (termed the “blocking effect” since Fe blocks further electron donation to the site) of the oxygen-evolving complex (OEC) in Mn-depleted, photosystem II (PSII) membrane fragments (Semin et al. in Biochemistry 41:5854, 2002). Furthermore, Fe(II) cations can substitute for 1 or 2Mn cations (pH dependent) in Ca-depleted PSII membranes (Semin et al. in Journal of Bioenergetics and Biomembranes 48:227, 2016; Semin et al. in Journal of Photochemistry and Photobiology B 178:192, 2018). In the current study, we examined the effect of Ca2+ cations on the interaction of Fe(II) ions with Mn-depleted [PSII(-Mn)] and Ca-depleted [PSII(-Ca)] photosystem II membranes. We found that Ca2+ cations (about 50 mM) inhibit the light-dependent oxidation of Fe(II) (5 µM) by about 25% in PSII(-Mn) membranes, whereas inhibition of the blocking process is greater at about 40%. Blocking of the HA site by Fe cations also decreases the rate of charge recombination between QA? and YZ?+ from t1/2?=?30 ms to 46 ms. However, Ca2+ does not affect the rate during the blocking process. An Fe(II) cation (20 µM) replaces 1Mn cation in the Mn4CaO5 catalytic cluster of PSII(-Ca) membranes at pH 5.7 but 2 Mn cations at pH 6.5. In the presence of Ca2+ (10 mM) during the substitution process, Fe(II) is not able to extract Mn at pH 5.7 and extracts only 1Mn at pH 6.5 (instead of two without Ca2+). Measurements of fluorescence induction kinetics support these observations. Inhibition of Mn substitution with Fe(II) cations in the OEC only occurs with Ca2+ and Sr2+ cations, which are also able to restore oxygen evolution in PSII(-Ca) samples. Nonactive cations like La3+, Ni2+, Cd2+, and Mg2+ have no influence on the replacement of Mn with Fe. These results show that the location and/or ligand composition of one Mn cation in the Mn4CaO5 cluster is strongly affected by calcium depletion or rebinding and that bound calcium affects the redox potential of the extractable Mn4 cation in the OEC, making it resistant to reduction.

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