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1.
Mn2+ uptake in the chick chorioallantoic membrane, an embryonic epithelial tissue which transports Ca2+ in vivo was studied using electron paramagnetic resonance (EPR). Mn2+ was used as a paramagnetic analog for Ca2+, since there is evidence that Mn2+ is accumulated by the Ca2+ transport mechanism.After 1.5 h of uptake the EPR spectrum of the Mn2+ in the membrane indicated that 89 % of the Mn2+ was in a spin-exchange form, indicating close packing of Mn2+. The Mn2+ spacing was estimated from the line width to be about 4.7 Å. The remaining Mn2+ was very likely Mn2+ hexahydrate.At pH 7.4 the spin-exchange spectrum tended to broaden when uptake was inhibited, while at pH 5.0 the spin-exchange spectrum was completely abolished in the presence of inhibitors.The EPR spectrum of Mn2+ in the chorioallantoic membrane had a broader line width than that of Mn2+ in isolated mitochondria, suggesting that in this tissue mitochondria are not directly involved in divalent cation transport. These EPR studies support the concept that divalent cations are sequestered in high concentrations from the rest of the cell contents during transcellular active transport.  相似文献   

2.
The complex of photosystem II (PSII) had been prepared from spinach by treatment with Triton X-100. The PSII, which had been depleted of the extrinsic 17- and 23-kDa polypeptides, was obtained by exposing the solution to a high concentration of NaCl, and the complex of PSII-La3+ was prepared by treatment with LaCl3. The result indicated that La3+ could inhibit the oxygen-evolution activity of PSII by replacing the Ca2+. The local structural environment of La in PSII has been also studied by using extended X-ray absorption fine structure (EXAFS). The primary result of EXAFS indicated that La coordinated with eight oxygen and/or nitrogen atoms, with the distance of the La-O/N bond being 2.5 Å. In addition, La coordinated with four carbon atoms, with a distance of 3.5 Å in the second shell. In the third shell, La coordinated with two manganese atoms, with the distance of La-Mn bond being 4.49 Å, and it was also found that the La-Mn distance (4.49 Å) was longer than that of Ca-Mn (3.3 Å) (1) in PSII.  相似文献   

3.
Human prolidase, the enzyme responsible for the hydrolysis of the Xaa-Pro/Hyp peptide bonds, is a key player in the recycling of imino acids during the final stage of protein catabolism and extracellular matrix remodeling. Its metal active site composition corresponding to the maximal catalytic activity is still unknown, although prolidase function is of increasing interest due to the link with carcinogenesis and mutations in prolidase gene cause a severe connective tissue disorder. Here, using EPR and ICP-MS on human recombinant prolidase produced in Escherichia coli (hRecProl), the Mn(II) ion organized in a dinuclear Mn(II)–Mn(II) center was identified as the protein cofactor. Furthermore, thermal denaturation, CD/fluorescence spectroscopy and limited proteolysis revealed that the Mn(II) is required for the proper protein folding and that a protein conformational modification is needed in the transition from apo- to Mn(II)loaded-enzyme. The collected data provided a better knowledge of the human holo-prolidase and, although limited to the recombinant enzyme, the exact identity and organization of the metal cofactor as well as the conformational change required for activity were proven.  相似文献   

4.
In oxygenic photosynthesis, water is oxidized at a protein-cofactor complex comprising four Mn atoms and, presumably, one calcium. Using multilayers of Photosystem II membrane particles, we investigated the time course of the disassembly of the Mn complex initiated by a temperature jump from 25°C to 47°C and terminated by rapid cooling after distinct heating periods. We monitored polarographically the oxygen-evolution activity, the amount of the YDox radical and of released Mn2+ by EPR spectroscopy, and the structure of the Mn complex by x-ray absorption spectroscopy (XAS, EXAFS). Using a novel approach to analyze time-resolved EXAFS data, we identify three distinct phases of the disassembly process: (1) Loss of the oxygen-evolution activity and reduction of YDox occur simultaneously (k1 = 1.0 min−1). EXAFS spectra reveal the concomitant loss of an absorber-backscatterer interaction between heavy atoms separated by ~3.3 Å, possibly related to Ca release. (2) Subsequently, two Mn(III) or Mn(IV) ions seemingly separated by ~2.7 Å in the native complex are reduced to Mn(II) and released (k2 = 0.18 min−1). The x-ray absorption spectroscopy data is highly suggestive that the two unreleased Mn ions form a di-μ-oxo bridged Mn(III)2 complex. (3) Finally, the tightly-bound Mn2(μ-O)2 unit is slowly reduced and released (k3 = 0.014 min−1).  相似文献   

5.
Hu Y  Yang JP  Liu JS 《Luminescence》2012,27(5):437-440
Mn‐doped willemite (Zn2SiO4:Mn) green phosphor were synthesized by sol–gel technology. The effect of the addition of sodium, as in the composition Zn(1.92 – X) NaXMn0.08SiO4, on the emission behavior was studied. FT–IR and EPR results revealed that sodium ion is incorporated into the lattice and results in the formation of isolated Mn2+ and Mn–Mn pairs. The maximum emission intensity of the sample under ultraviolet (UV) excitation occurred at the sodium concentration of x = ~0.03. The green emission at about 525 nm is assigned to Mn2+–Mn2+ pair centres on nearest neighbour Zn sites. The highest intensity of the green emission for x = ~0.03 is well close to the highest concentration of the Mn2+–Mn2+ pair. Copyright © 2012 John Wiley & Sons, Ltd.  相似文献   

6.
《BBA》1986,850(2):324-332
The structure of the Mn complex in the oxygen-evolving system and its mechanistic relation to photosynthetic oxygen evolution are poorly understood, though many studies have established that membrane-bound Mn plays an active role. Recently established procedures for isolating oxygen-evolving subchloroplast Photosystem II (PS II) preparations and the discovery of a light-induced multiline EPR signal attributable to the S2 state of the O2-evolving complex have facilitated the preparation of samples well characterized in the S1 and S2 states. We have used extended X-ray absorption fine structure (EXAFS) spectroscopy to probe the ligand environment of Mn in PS II particles from spinach, and in this report we present our results. The essential feature of the EXAFS results are that at least two Mn atoms per PS II reaction center occur as a binuclear species with a metal-metal distance of approx. 2.7 Å, with low Z atoms, N or O, at a distance of approx. 1.75 Å and at approx. 1.98 Å, which are characteristic of bridging and terminal ligands. These results agree well with those derived from whole chloroplasts that provided the first evidence for a binuclear manganese complex (Kirby, J.A., Robertson, A.S., Smith, J.P., Thompson, A.C., Cooper, S.R. and Klein, M.P. (1981) J. Am. Chem. Soc. 103, 5529–5537).  相似文献   

7.
Mo K-edge X-ray absorption spectroscopy (XAS) has been used to probe the environment of Mo in dimethylsulfoxide (DMSO) reductase from Rhodobacter capsulatus in concert with protein crystallographic studies. The oxidised (MoVI) protein has been investigated in solution at 77?K; the Mo K-edge position (20006.4?eV) is consistent with the presence of MoVI and, in agreement with the protein crystallographic results, the extended X-ray absorption fine structure (EXAFS) is also consistent with a seven-coordinate site. The site is composed of one oxo-group (Mo=O 1.71?Å), four S atoms (considered to arise from the dithiolene groups of the two molybdopterins, two at 2.32?Å and two at 2.47?Å, and two O atoms, one at 1.92?Å (considered to be H-bonded to Trp 116) and one at 2.27?Å (considered to arise from Ser 147). The Mo K-edge XAS recorded for single crystals of oxidised (MoVI) DMSO reductase at 77?K showed a close correspondence to the data for the frozen solution but had an inferior signal:noise ratio. The dithionite-reduced form of the enzyme and a unique form of the enzyme produced by the addition of dimethylsulfide (DMS) to the oxidised (MoVI) enzyme have essentially identical energies for the Mo K-edge, at 20004.4?eV and 20004.5?eV, respectively; these values, together with the lack of a significant presence of MoV in the samples as monitored by EPR spectroscopy, are taken to indicate the presence of MoIV. For the dithionite-reduced sample, the Mo K-edge EXAFS indicates a coordination environment for Mo of two O atoms, one at 2.05?Å and one at 2.51?Å, and four S atoms at 2.36?Å. The coordination environment of the Mo in the DMS-reduced form of the enzyme involves three O atoms, one at 1.69?Å, one at 1.91?Å and one at 2.11?Å, plus four S atoms, two at 2.28?Å and two at 2.37?Å. The EXAFS and the protein crystallographic results for the DMS-reduced form of the enzyme are consistent with the formation of the substrate, DMSO, bound to MoIV with an Mo-O bond of length 1.92?Å.  相似文献   

8.
《FEBS letters》1985,189(2):355-360
Inhibition of oxygen evolution in photosystem II membrane fragments from pea chloroplasts by washing with Zn2+ causes appearance of the EPR signal of Mn(H2O)62+. This Mn2+ remains associated with the membrane fraction. Release of Mn2+ into the medium was correlated with the amount of the 23 kDa protein removed from the membrane. This suggests that this protein may function as a ‘gate’ to an aqueous compartment into which Mn2+ is released. Inhibition by Zn2+ correlated with the release of 1 Mn2+ per reaction centre, out of a total stoichiometry of 4 Mn atoms per reaction centre. By comparing the release of Mn following Zn-treatment of NaCI or CaC12 washed membranes, it is concluded that the 33 kDa protein is involved in binding of 2 Mn.  相似文献   

9.
Treatment of thylakoid suspensions with Zn2+ causes the appearance of an EPR signal due to Mn2+. The size of the signal was linearly correlated with the inhibition of oxygen evolution. Full inhibition appeared to correspond to the release of 2 Mn atoms/reaction centre of photosystem II. The released Mn2+ remained associated with the chloroplast pellet on centrifugation and took several hours to equilibrate with the surrounding medium. The sequestered Mn2+ does not appear to be in the thylakoid interior but in a more restricted hydrophilic compartment.  相似文献   

10.
A detailed analysis of the EPR signatures at X-band and Q-band of an enzyme (SoxB) involved in sulfur oxidation from Paracoccus pantotrophus is presented. EPR spectra are attributed to an exchange-coupled dimanganese Mn2(II,II) complex. An antiferromagnetic exchange interaction of J=?7.0 (±1) cm?1 (H=?2JS 1 S 2 ) is evidenced by a careful examination of the temperature dependence of the EPR spectra. The spin Hamiltonian parameters for a total spin of S T =1, 2 and 3 are obtained and an inter-manganese distance of 3.4 (±0.1) Å is estimated. The comparison with exchange coupling and inter-manganese distance data of other dimanganese proteins and model compounds leads to a tentative assignment of the Mn bridging ligands to bis(μ-hydroxo) (μ-carboxylato).  相似文献   

11.
Three new chiral ligands bearing an O,O′,N donor set (OmethoxyOhydroxyNpyridine) were synthesised and coordinated to FeIII, FeII, NiII, CuII and ZnII to yield complexes with the general formula [M(OON)Clx]y. While the pyridine N and the hydroxy O atoms coordinate strongly to all applied metal ions, the methoxy donor seems not to be involved in coordination, although some evidence for a weak interaction between OMe and the ZnII were found in NMR spectra. In the bidentate O′,N coordination mode the new ligands exhibit several coordination geometries as analysed in the solid compounds by XRD, EXAFS and EPR and in solution by UV-Vis absorption, cyclic voltammetry, EXAFS, EPR or NMR spectroscopy.  相似文献   

12.
《Inorganica chimica acta》2001,312(1-2):245-248
The structure of the title compound has been determined by X-ray diffraction at 190 K. The complex has an all trans configuration with an elongated tetragonally distorted octahedral CuO6 chromophore. The elongated axis corresponding to the trans-Cu–O(ether) bonds. The ligand molecules are bidentate via the carboxyl and the 3-ether O atoms; the 6-ether O atoms are not coordinated and are remote from the Cu centres. The bond lengths to the Cu centres are Cu–O(ether) 2.355 Å, Cu–O(Carboxyl) 1.933 Å and Cu–OH2 1.995 Å.The EPR spectrum of both the powder and frozen solution forms is typical of a rhombic system with a dx2y21 electronic configuration. There were no significant differences in spectra recorded over the temperature range 77 K to room temperature. These results are discussed in relation to earlier published results on closely related oxa-carboxyl complexes.  相似文献   

13.
The effect of high Mn2+ content on Centaurium pulchellum seed germination has been investigated. Seeds containing extremely high Mn2+ content were produced by culturing single-node flowering explants for 2 months in the MS-media, supplemented with Mn in concentrations ranging from 1 to 10,000 μM. Although the seeds displayed the capacity to accumulate high amount of Mn, their germination was undisturbed. EPR spectroscopy was used to measure the ratio of free (aqueous) Mn to bound Mn and it was found that over 97% of total Mn was in the bound form. With elevating the external Mn supply, seed Mn concentration also increased, but the proportion of free Mn2+ fraction decreased from 3% in the control (1 μM Mn) to 0.35% and 0.15% in high Mn supply (1000 μM and 10,000 μM, respectively). These results suggest that an elevation of internal Mn concentration in seeds is associated with increased Mn binding pools, hence Mn remains bound during germination. Consequently, the action of potentially harmful Mn2+ ions, which may generate ROS and affect seed viability, is alleviated.  相似文献   

14.
Rita Khanna  S. Rajan  H.S. Gutowsky 《BBA》1983,725(1):10-18
Measurements were made of the water proton relaxation rate (T?12 = R2), electron spin resonance (ESR) six-line signal of ‘free’ Mn2+, and O2-evolution activity in thylakoid membranes from pea leaves. The main results are: (1) Aging of thylakoids at 35°C causes a parallel decrease in O2-evolution activity, in R2 and in the content of bound Mn, suggesting that R2 may be related to the loosely bound Mn involved in O2 evolution. (2) Treatment of thylakoids with tetraphenylboron (TPB) at [TPB] > 2 mM produces a 2-fold increase in R2, without release of Mn2+. The titration curve exhibits three sharp end points. The first end point occurs at a [TPB][chlorophyll] of 1.25, at which the O2 evolution is completely inhibited. (3) Treatment of thylakoids with NH2OH also increases R2 by nearly 2-fold, either by the reduction of the higher oxidation states of Mn to Mn2+ and / or by exposing the Mn to solvent protons. Also, progressive release of bound Mn occurs at [NH2OH] ≥ 1 mM as shown by an increase increase in the Mn2+ ESR signal and a decrease in R2. (4) Addition of H2O2 (0.1–1.0%) to thylakoids causes an enhancement of R2 similar to that by NH2OH, but without the release of Mn2+. (5) Heat treatment of thylakoids at 40–50°C releases Mn2+ and increases R2. Conversely, pH values of 7 to 4 release Mn2+ without changing R2 while pH values of 7–9 increase R2 without releasing Mn2+. Thus, both high and low pH values as well as the heat treatment cause structural changes enhancing the relaxivity of the bound Mn or of other paramagnetic species.  相似文献   

15.
《Inorganica chimica acta》1988,142(2):333-336
The crystal and molecular structures of the title compounds have been determined from single crystal X-ray diffraction. The two complexes crystallize in the orthorhombic space group Pna21 with Z = 4. Lattice parameters are: a = 10.504(2) [10.522(2)], 1b = 16.816(4) [16.927(3)] and c = 18.931(4) [18.969(3)] Å. The two crystals are isomorphous. The structures were solved by Patterson and Fourier techniques and refined by least-squares techniques to R = 0.0430 for 1508 reflections. The Nd3+. ion is eight-coordinate, being bonded to five carbons of the cyclopentadiene ring, to four chloride atoms and to the one oxygen atom of the THF ring. The NdC distances are in the range 2.67–2.85 Å (average: 2.77 Å) and Nd-Cl distances are in the range 2.76–2.80 Å (average: 2.78 Å). The Nd-O distance is 2.52 A. The Li+ ion is four-coordinate, being bonded to the two chloride atoms and to the two oxygen atoms of the two THF rings. The other Li+ ion is the same as the above. The Li-Cl distances are in the range 2.17– 2.55 Å (average: 2.35 A) and LiO distances are in the range 1.89–1.98 Å (average: 1.91 Å). The Nd atom and the two Li atoms are bridged asymmetrically by the chloride ions, respectively.  相似文献   

16.
Proton and phosphorus-31 nuclear spin–lattice relaxation times T1 and spin–spin relaxation times T2 have been measured on the single-stranded polyriboadenylic acid [poly(A)]–Mn2+ system in a neutral D2O solution in the temperature range 10°–90°C at 100 and 40.5 MHz, respectively, with the Fourier transform nmr method. Minimum values of T1 have been found for all these nuclei, which have enabled the exact estimation of apparent distances from Mn2+ to H2, H8, H1′, and the phosphorus nucleus to be 4.7, 4.1, 5.2, and 3.0 Å, respectively. The electron spin of Mn2+ penetrates into the phosphorus nucleus, giving 31P hyperfine coupling of more than 106 Hz. Evidence of penetration of the electron spin into H8 and H2 is also obtained, suggesting direct coordination of nitrogen atoms of the adenine ring to the Mn2+ Ion. Combined with the result from proton relaxation enhancement of water, it is concluded that every Mn2+ ion added is bound directly to two phosphate groups with a Mn2+–phosphorus distance of 3.3 Å, while a part of the Mn2+ ions are simultaneouly bound to the adenine ring. It is estimated that 39 ± 13% and 13 ± 5% of Mn2+ are coordinated by N7 and N3 (or N1), respectively. The motional freedom of poly(A) in the environment of the Mn2+ binding site has been found to be quenched to the extent that the rotational motion becomes several times slower than that of the corresponding Mn2+–free poly(A). The activation energies for the molecular motion are, however, practically unchanged from those for Mn2+–free poly(A), and are found to be 8.3, 8.5, 6.1, and 8.7 kcal/mol for H8, H2, H1′, and phosphorus, respectively. T2 of phosphorus is determined by the dissociation rate (k?1) of Mn2+ from the phosphate group for the whole temperature range studied with activation enthalpy of 6.5 kcal/mol. The dissociation rates of Mn2+ from the adenine ring are also estimated from proton T2 values below 50°C.  相似文献   

17.
Myoglobin of Aplysia brasiliana (MbApB) has been recently purified and characterized and it was shown that the amino acid content is quite different from other myoglobins. A large number of aromatic residues was observed together with the existence of a unique histidine at the proximal heme position. Because of the numerous differences in the amino acid sequence between MbApB and whale myoglobin, it was interesting to investigate the interaction of metal ions like Cu2+ and Mn2+ with MbApB. In the present work Cu2+ complexes with Met-MbApB were studied and show a pH transition between different forms of coordination as revealed by EPR measurements. At high pH the EPR spectrum shows the coordination of the metal to at least four nitrogens from ϵ-NH3 lysine residues. At lower pH in the range 6.0–9.0 the copper binding site shows a pK change of some of the residues involved in metal coordination. Addition of one equivalent Cu2+ per protein does not alter the iron EPR signal. The manganese ion has one binding site in MbApB and a binding constant Ka = ( 11.5 ± 0.8) 103M−1. The binding of Cu2+ to MbApB is stronger than Mn2+, KaCu2+ >KaMn2+.  相似文献   

18.
The cyanobacterium Oscillatoria terebriformis was shown to exhibit resistance to high manganese concentrations, remaining viable at 2.5 mM MnCl2 in the medium. Cyanobacterial cells were capable of considerable manganese consumption from the medium. The dynamics of Mn sorption by the cells were the same in all experimental variants, independent of the manganese concentration. Manganese concentration in the biomass peaked after 2–3 days and depended on Mn2+ concentration in the medium and on the amount of biomass introduced. In the case of O. terebriformis, manganese removed from the medium may be subdivided into Mn absorbed by the cell, Mn bound to the cell wall, Mn absorbed by the glycocalix, and chemically precipitated Mn. Of the total 21.25 ± 1.0 mg of consumed manganese, biological absorption and chemical precipitation were responsible for 11.78 ± 0.98 and 9.2 ± 0.8 mg, respectively. In the presence of cyanobacteria, Mn removal from the medium was 2.28 times higher than in the control. This process depended considerably on Mn sorption by exopolysaccharides. At 1.3 mM Mn2+, a lamellar mat was formed with interlayers of manganese carbonate.  相似文献   

19.
The interactions of Mn2+ and Co2+ with glucose isomerases from three microbial sources have been studied using various direct physical methods. Co2+ was found to activate each enzyme, although the degree of activation varied significantly for enzymes from different organisms. EPR spectroscopy measurements revealed that dissimilarities in the coordination sphere of enzyme-bound Mn2+ accompanied the differences in enzyme activity. Variations in the EPR spectra of a nitroxide spin label coupled to two of the three isomerases, possibly near their active sites, were also observed. In no case was the EPR spectrum influenced by Co2+ addition, a result discordant with the hypothesis that Co2+ activates the enzyme by inducing a conformational change. The proximal biochemical environment of enzymebound Co2+ was also examined using EXAFS spectroscopy. This method showed that glucose causes notable changes in the ligand environment of the enzyme-bound metal, suggesting the formation of an enzyme-metal-substrate bridge complex. The significance of these results relative to possible reaction mechanisms is discussed.  相似文献   

20.
Isopentenyl pyrophosphate isomerase (EC 5.3.3.2) from pig liver has been purified 197-fold. The preparation was estimated to contain less than 10% of contaminating protein. The molecular weight determined by gel filtration was 82,500 ± 3,000 and the isoelectric point from isoelectric focusing was in the range 6.0–6.2. N-terminal analysis showed the presence of both leucine and proline. The pH optimum of the enzyme preparation was 6.3. After dialysis against EDTA, activity was restored by either Mn2+ or Mg2+, the former being more effective. At the optimum pH and concentration of Mn2+, Km and V were 2.7 μm and 6.7 μmol min?1 mg?1, respectively. The enzyme was partially inhibited by a variety of terpene mono- and pyrophosphate esters, by inorganic phosphate ions, and by acetate ions; essentially complete inhibition by sulfhydryl-blocking reagents was observed. ATP partially inhibited, the degree of inhibition showing a sigmoid dependence on ATP concentration. Monothiols and dithiothreitol activated the enzyme, as did mevalonic acid.  相似文献   

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