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1.
The large enhancement of the green luminescence of terbium ion which occurs on binding to porcine and bovine trypsins and to bovine α-chymotrypsin has been used to study the calcium binding sites of these enzymes. Excitation spectra, taken at low protein concentrations to minimize absorption effects, demonstrate that in each case, energy transfer occurs between the side chain of a tryptophan residue and bound Tb3+. Association constants for the binding of Tb3+ to the single binding site on each of the three proteins have been measured at 25 °C and pH 6.6. Ca2+ ions compete with Tb3+ for the single binding site, and association constants for Ca2+ were determined by Tb3+ displacement. The ratio of binding strengths of Ca2+ to α-chymotrypsin, bovine trypsin, porcine trypsin, and elastase is 1:12:24:23. Addition of Tb3+ to the homologous bacterial enzyme α-lytic protease caused no luminescence enhancement.  相似文献   

2.
BaO‐B2O3‐P2O5 glasses doped with a fixed concentration of Tb3+ ions and varying concentrations of Al2O3 were synthesized, and the influence of the Al3+ ion concentration on the luminescence efficiency of the green emission of Tb3+ ions was investigated. The optical absorption, excitation, luminescence spectra and fluorescence decay curves of these glasses were recorded at ambient temperature. The emission spectra of terbium ions when excited at 393 nm exhibited two main groups of bands, corresponding to 5D3 → 7Fj (blue region) and 5D4 → 7Fj (green region). From these spectra, the radiative parameters, viz., spontaneous emission probability A, total emission probability AT, radiative lifetime τ and fluorescent branching ratio β, of different transitions originating from the 5D4 level of Tb3+ ions were evaluated based on the Judd‐Ofelt theory. A clear increase in the quantum efficiency and luminescence of the green emission of Tb3+ ions corresponding to 5D4 → 7F5 transition is observed with increases in the concentration of Al2O3 up to 3.0 mol%. The improvement in emission is attributed to the de‐clustering of terbium ions by Al3+ ions and also to the possible admixing of wave functions of opposite parities. Copyright © 2016 John Wiley & Sons, Ltd.  相似文献   

3.
Ciliate Euplotes octocarinatus centrin (EoCen) is a member of the EF-hand superfamily of calcium-binding proteins. It has been proven, using Tb3+ as a fluorescence probe, that EoCen has four calcium-binding sites. The sensitized emission arises from nonradiative energy transfer between the three tyrosine residues (Tyr46, Tyr72, and Tyr79) of the N-terminal half and the bound Tb3+ ions. To determine the most critical of the three tyrosine residues for the process of fluorescence resonance energy transfer, six mutants of the N-terminal domain of EoCen, which contain one (N-Tyr46/N-Tyr72/N-Tyr79) or two (N-Y46F/N-Y72F/N-Y79F) tyrosine residues, were obtained by site-directed mutagenesis. The aromatic residue-sensitized Tb3+ fluorescence of N-Y79F was most affected, displaying a 50% reduction compared with wild-type N-EoCen. Among the tyrosines, Tyr79 is the shortest mean distance from the protein-bound Tb3+ (at sites I/II), as calculated via the Förster mechanism. The steady-state and time-resolved fluorescence parameters of the wild-type N-EoCen and the three double mutants suggest that Tyr79, which exists in a hydrophobic environment, has the highest quantum yield and a relatively long average lifetime. The decay of Tyr79 is the least heterogeneous among the three tyrosine residues. In addition, molecular modeling shows that a critical hydrogen bond is formed between the 4-hydroxyl group of Tyr79 and the oxygen from the side chains of the residue Asn39. Kinetic experiments on tyrosine and Tb3+ fluorescence demonstrate that tyrosine fluorescence quenching is largely due to the self-assembly of EoCen, and that the quenching degrees of the mutants differ. Resonance light scattering and crosslinking analysis carried out on the full-length single mutants (Y46F, Y72F, and Y79F) showed that Tyr79 also plays the most important role in the Tb3+-dependent self-assembly of EoCen among the three tyrosines.  相似文献   

4.
The fluorescence emission intensity of terbium is enhanced upon the binding of Tb3+ to cultured mouse spinal cord and dorsal root ganglion neurons, via nonradiative resonant energy transfer from membrane proteins. The relative fluorescence intensities of Tb3+ bound to dorsal root ganglion neurons were considerably greater than that of Tb3+ bound to large multipolar spinal cord neurons. The cell bodies of the dorsal root ganglion neurons were completely covered in a dense fluorescent blanket, whereas the fluorescence from the spinal cord soma presented a discontinuous pattern. The neurites of the spinal cord neuron were speckled with bright patches of Tb3+ fluorescence. A high concentration of Ca2+ reduced the relative fluorescence intensity of the Tb3+ -neuron complex. It is suggested that Tb3+ binds to Ca2+ -binding sites on the surface membrane of neurons.  相似文献   

5.
Seven novel pyrazolone derivatives were synthesized and characterized by 1H NMR and 13C NMR spectra, mass spectra, infrared spectra and elemental analysis. Their terbium complexes were prepared and characterized by elemental analysis, EDTA titrimetric analysis, UV/vis spectra, infrared spectra and molar conductivity, as well as thermal analysis. The fluorescence properties and fluorescence quantum yields of the complexes were investigated at room temperature. The results indicated that pyrazolone derivatives had good energy‐transfer efficiency for the terbium ion. All the terbium complexes emitted green fluorescence characteristic of terbium ions, possessed strong fluorescence intensity, and showed relatively high fluorescence quantum yields. Cyclic voltammograms of the terbium complexes were studied and the highest occupied molecular orbital (HOMO) and lowest occupied molecular orbital (LUMO) energy levels of these complexes were estimated. Copyright © 2014 John Wiley & Sons, Ltd.  相似文献   

6.
Two complexes of Tb3+, Gd3+/Tb3+ and one heteronuclear crystal Gd3+/Tb3+ with phenoxyacetic acid (HPOA) and 2,4,6‐tris‐(2‐pyridyl)‐s–triazine (TPTZ) have been synthesized. Elemental analysis, rare earth coordination titration, inductively coupled plasma atomic emission spectrometry (ICP‐AES) and thermogravimetric analysis‐differential scanning calorimetry (TG‐DSC) analysis show that the two complexes are Tb2(POA)6(TPTZ)2·6H2O and TbGd(POA)6(TPTZ)2·6H2O, respectively. The crystal structure of TbGd(POA)6(TPTZ)2·2CH3OH was determined using single‐crystal X‐ray diffraction. The monocrystal belongs to the triclinic system with the P‐1 space group. In particular, each metal ion is coordinately bonded to three nitrogen atoms of one TPTZ and seven oxygen atoms of three phenoxyacetic ions. Furthermore, there exist two coordinate forms between C6H5OCH2COO and the metal ions in the crystal. One is a chelating bidentate, the other is chelating and bridge coordinating. Fluorescence determination shows that the two complexes possess strong fluorescence emissions. Furthermore, the fluorescence intensity of the Gd3+/Tb3+ complex is much stronger than that of the undoped complex, which may result from a decrease in the concentration quench of Tb3+ ions, and intramolecular energy transfer from the ligands coordinated with Gd3+ ions to Tb3+ ions. Copyright © 2015 John Wiley & Sons, Ltd.  相似文献   

7.
Carp muscle calcium binding parvalbumin, crystallized in 2.9 m-ammonium sulfate, can bind two Tb3+ ions, which displace the two Ca2+ ions normally present. The Ca2+ co-ordinated in the loop between the E and the F α-helices is displaced at low Tb3+ concentrations; whereas the Ca2+ at the CD site is replaced only at higher Tb3+ concentration. There is not a third Tb3+ site as had been suggested in interpretations of Tb3+ fluorescence experiments performed without ammonium sulfate. A third electron density peak in the difference Fourier maps is tentatively assigned to a sulfate ion co-ordinating the EF site Tb3+ ion.  相似文献   

8.
A series of SrMoO4:Sm3+,Tb3+,Na+ phosphors was synthesized using a high‐temperature solid‐state reaction method in air. On excitation at 290 nm, SrMoO4:Sm3+,Tb3+ phosphor emitted light that varied systematically from green to reddish‐orange on changing the Sm3+ and Tb3+ ion concentrations. The emission intensities of SrMoO4:Sm3+ and SrMoO4:Sm3+,Tb3+ phosphors were increased two to four times due to charge compensation when Na+ was added as a charge compensator. The luminescence mechanism and energy transfer could be explained using energy‐level diagrams of the MoO42– group, Sm3+ and Tb3+ ions. SrMoO4:Sm3+,Tb3+,Na+ could be used as reddish‐orange phosphor in white light‐emitting diodes (LEDs) based on an ~ 405 nm near‐UV LED chip. This research is helpful in adjusting and improving the luminescence properties of other phosphors. Copyright © 2015 John Wiley & Sons, Ltd.  相似文献   

9.
Two novel FRET-pairs: Tb3+-binding peptide-DsRed2 and Tb3+-binding peptide-TagRFP have been produced based on the terbium-binding peptide and the red fluorescent proteins DsRed2 and TagRFP. Two induction-resonance energy transfer processes in both FRET-pairs have been registered, from tryptophan of the terbium-binding peptide to Tb3+ and from sensitized Tb3+ to the acceptor, the chromophore, DsRed2 or TagRFP. The lifetimes of terbium in the presence and absence of the acceptor have been determined. It has been shown that the lifetime of Tb3+ in the presence of 150 mM NaCl decreases in both FRET-pairs. The efficiency of fluorescent resonance transfer from Tb3+ to the acceptor proteins is 28 and 35% for Tb3+-binding peptide-DsRed2 and Tb3+-binding peptide-TagRFP, respectively.  相似文献   

10.
Using small-angle x-ray diffraction from centrifugally oriented acetylcholine receptor (AChR) enriched membranes coupled with anomalous scattering from terbium ions (Tb3+) titrated into presumed Ca2+ binding sites, we have mapped the distribution of Tb3+ perpendicular to the membrane plane using a heavy atom refinement algorithm. We have compared the distribution of Tb3+ in the closed resting state with that in the carbamylcholine-desensitized state. In the closed resting state we find 45 Tb3+ ions distributed in 10 narrow peaks perpendicular to the membrane plane. Applying the same refinement procedure to the data from carbamylcholine desensitized AChR we find 18 fewer Tb3+ ions in eight peaks, and slight rearrangements of Tb3+ density in the peaks near the ends of the AChR ion channel pore. These agonist dependent changes in the Tb3+ stoichiometry and distribution suggest a likely role for multivalent cations in stabilizing the different functional states of the AChR, and the changes in the Tb3+ distribution at the two ends of the pore suggest a potential role for multivalent cations in the gating of the ion channel.  相似文献   

11.
A terbium‐sensitized spectrofluorimetric method has been developed for determination of catecholamines such as norepinephrine (NE), epinephrine (EP) and dopamine (DA), using sodium dodecyl benzene sulphonate (SDBS). Fluorescence sensitization of terbium ions (Tb3+) by complexation with catecholamines in the presence of SDBS was observed. The fluorescence intensities of the Tb3+–catecholamine complexes were highly enhanced by introducing SDBS with an emission maximum at 545 nm after excitation at 290 nm. The conditions for the complex formation of Tb3+–catecholamine were investigated systematically and optimized to determine catecholamines in a serum sample. Under the optimum conditions, the fluorescence intensities of the Tb3+–catecholamine complexes were increased linearly with the concentration of NE, EP and DA over the ranges 2.5 × 10–10–1.0 × 10–8, 2.5 × 10–10–1.0 × 10–8 and 2.5 × 10–9–1.0 × 10–7 g/mL with correlation coefficients of 0.999, 0.999 and 0.9996, respectively. The limits of detection (3δ) of NE, EP and DA were found to be 4.6 × 10–11, 7.8 × 10–11 and 8.38 × 10–10 g/mL, respectively. Precision of the method was tested at the concentration level of 1.2 × 10?7 g/mL for five replicate measurements of NE, EP and DA, giving relative standard deviations (RSDs) of 1.41%, 1.23% and 1.89%, respectively. The interaction mechanism of the Tb3+–catecholamine complexes system was investigated and presented with ultraviolet absorption spectra. The proposed method has been applied for the quantitative determination of NE, EP and DA in a spiked serum sample and a pharmaceutical preparation sample. Copyright © 2011 John Wiley & Sons, Ltd.  相似文献   

12.
Single crystals of KCl doped with Ce3+,Tb3+ were grown using the Bridgeman–Stockbarger technique. Thermoluminescence (TL), optical absorption, photoluminescence (PL), photo‐stimulated luminescence (PSL), and thermal‐stimulated luminescence (TSL) properties were studied after γ‐ray irradiation at room temperature. The glow curve of the γ‐ray‐irradiated crystal exhibits three peaks at 420, 470 and 525 K. F‐Light bleaching (560 nm) leads to a drastic change in the TL glow curve. The optical absorption measurements indicate that F‐ and V‐centres are formed in the crystal during γ‐ray irradiation. It was attempted to incorporate a broad band of cerium activator into the narrow band of terbium in the KCl host without a reduction in the emission intensity. Cerium co‐doped KCl:Tb crystals showed broad band emission due to the d–f transition of cerium and a reduction in the intensity of the emission peak due to 5D37Fj (j = 3, 4) transition of terbium, when excited at 330 nm. These results support that energy transfer occurs from cerium to terbium in the KCl host. Co‐doping Ce3+ ions greatly intensified the excitation peak at 339 nm for the emission at 400 nm of Tb3+. The emission due to Tb3+ ions was confirmed by PSL and TSL spectra. Copyright © 2015 John Wiley & Sons, Ltd.  相似文献   

13.
Diffusion-enhanced fluorescence energy transfer was used to study the structure of photoreceptor membranes from bovine retinal rod outer segments. The fluorescent energy donor was Tb3+ chelated to dipicolinate and the acceptor was the 11-cis retinal chromophore of rhodopsin in vesicles made from disc membranes. The rapid-diffusion limit for energy transfer was attained in these experiments because of the long excited state lifetime of the terbium donor (~2 ms). Under these conditions, energy transfer is very sensitive to a, the distance of closest approach between the donor and acceptor (Thomas et al., 1978). Vesicles containing terbium dipicolinate in their inner aqueous space were prepared by sonicating disc membranes in the presence of this chelate and chromatographing this mixture on a gel filtration column. The sidedness of rhodopsin in these vesicles was the same as in native disc membranes. The transfer efficiency from terbium to retinal in this sample was 43%. For an R0 value of 46.7 Å and an average vesicle diameter of 650 Å, this corresponds to an a value of 22 Å from the inner aqueous space of the vesicle. The distance of closest approach from the external aqueous space, determined by adding terbium dipicolinate to a suspension of already formed vesicles, was found to be 28 Å. These values of a show that the retinal chromophore is far from both aqueous surfaces of the disc membrane. Hence, the transverse location of the retinal chromophore is near the center of the hydrophobic core of the disc membrane. These findings suggest that conformational changes induced by photoisomerization are transmitted through a distance of at least 20 Å within rhodopsin to trigger subsequent events in visual excitation.  相似文献   

14.
The interaction of a number of lanthanide ions (namely terbium, praseodymium, erbium, lanthanum, gadolinium and europium) with Panulirus interruptus hemocyanin has been studied.Results from O2-binding experiments indicate that all these ions may substitute for calcium as allosteric effectors of hemocyanin. Addition of the lanthanides to deoxygenated Panulirus hemocyanin saturated with Tb3+ results in a quenching of the terbium luminescence. The highly efficient quenching observed in the case of Eu3+ may indicate energy-transfer between Tb3+ and Eu3+. Since energy-transfer between lanthanides is only effective over very short distances, the data suggest that some of the cation binding sites of Panulirus hemocyanin are clustered.  相似文献   

15.
Anticoagulation factor I (ACF I) isolated from the venom of Agkistrodon acutus is an activated coagulation factor X-binding protein with marked anticoagulant activity. Present studies show that holo-ACF I assumes a compactly folded structure in the range of pH 5–6, in which the most interior Trp residues and quenchers are adjacent. Tb3+ ions can completely replace both Ca2+ ions in holo-ACF I, as determined by equilibrium dialysis. Although the two Tb3+ ions in Tb3+-ACF I have slightly different luminescence efficiencies, both have similar quenching effects on the intrinsic fluorescence, suggesting that probably there are same numbers of Trp residues close to both Tb3+-binding sites. Two Tb3+-binding sites with similar apparent Tb3+ association constant values, (1.69 ± 0.02) × 107 M–1 and (1.42 ± 0.01) × 107 M–1, respectively, were further identified through Tb3+ fluorescence titration. In addition, it has been confirmed from the titration of holo-ACF I and Tb3+-ACF I with NBS that only interior Trp residues are involved in the energy transfer to Tb3+ ions and that all accessible Trp residues located in the surface of holo-ACF I have similar affinity to NBS, while those located in the surface of Tb3+-ACF I have two different kinds of affinity to NBS, which strongly suggests a conformational change of holo-ACF I upon substitution of Tb3+ for Ca2+. The results show that although the Tb3+-altered conformation of ACF I cannot support the binding of Tb3+-ACF I with FXa, determined by nondenaturing PAGE, Tb3+ ions are effective and useful fluorescence probes to analyze the structures and properties of Ca2+-binding sites in ACF I.  相似文献   

16.
17.
A rapid, simple, selective and precise fluorimetric method was developed and validated for determination of a selective xanthine oxidase inhibitor; febuxostat (FBX) in pharmaceutical formulations and in human plasma. The proposed method is based on quenching effect of FBX on the fluorescence intensity of terbium (Tb3+) through fluorescence resonance energy transfer (FRET) from Tb3+ to FBX. The formed complex was measured at λex. 320 nm/λem. 490 nm against a reagent blank. Fluorescence intensity of Tb3+ was diminished when FBX was added. A linear relationship between the fluorescence quenching value of the formed complex and the concentration of FBX was investigated. The reaction conditions and the fluorescence spectral properties of the complex have been studied. The linearity range of the developed method was 1.0–16.0 μg/ml. The suggested method was applied successfully for the estimation of FBX in bulk powder, dosage forms and spiked plasma samples with excellent recoveries (96.79–98.89%). In addition, the developed method has been successfully applied for determination of FBX in real plasma samples collected from healthy volunteers with good recoveries (82.06–85.65%). All obtained results of the developed method were statistically analyzed and validated according to ICH (International Conference on Harmonization) guidelines.  相似文献   

18.
Eu2+ and Tb3+ singly doped and co‐doped LaAl11O18 phosphors were prepared by a combustion method using urea as a fuel. The phase structure and photoluminescence (PL) properties of the prepared phosphors were characterized by powder X‐ray diffraction (XRD), scanning electron microscopy (SEM), and photoluminescence excitation and emission spectra. When the content of Eu2+ was fixed at 0.01, the emission chromaticity coordinates could be adjusted from blue to green region by tuning the contents of Tb3+ ions from 0.01 to 0.03 through an energy transfer (ET) process. The fluorescence data collected from the samples with different contents of Tb3+ into LaAl11O18: Eu, show the enhanced green emission at 545 nm associated with 5D47F5 transitions of Tb3+. The enhancement was attributed to ET from Eu2+ to Tb3+, and therefore Eu2+ ion acts as a sensitizer (an energy donor) while Tb3+ ion as an activator. The ET from Eu2+ to Tb3+ is performed through dipole–dipole interaction. The ET efficiency and critical distance were also calculated. The present Eu2+–Tb3+ co‐doped LaAl11O18 phosphor will have potential application for UV convertible white light‐emitting diodes. Copyright © 2015 John Wiley & Sons, Ltd.  相似文献   

19.
Eight novel 1,2,4‐triazolo[3,4‐b]‐1,3,4‐thiadiazol derivatives have been designed and synthesized, and their corresponding Tb3+ complexes were also prepared successfully. The fluorescence properties and fluorescence quantum yields of the target complexes were investigated, the results showed that the ligands were an efficient sensitizer for Tb3+ luminescence, and the target complexes exhibited characteristic fluorescence emissions of Tb3+ ion. The fluorescence intensity of the complex substituted by chlorine was stronger than that of other complexes. The substituents' nature has a great effect upon the electrochemical properties of the target complexes. The results showed that the introduction of the electron‐withdrawing groups tended to decrease the oxidation potential and highest occupied molecular orbital energy levels of the target Tb3+ complexes; however, introduction of the electron‐donating groups can increase the corresponding complexes' oxidation potential and highest occupied molecular orbital energy levels. Copyright © 2014 John Wiley & Sons, Ltd.  相似文献   

20.
Calcium ion-induced fusion events in suspensions of large unilamellar phosphatidylserine (PS) liposomes were monitored by fluorescence methods. Mixing of vesicle contents was studied by measuring the increase in terbium emission intensity due to formation of a complex between Tb3+ ions and dipicolinic acid trapped in the liposomes. Lipid redistribution was determined with the aid of the resonance transfer of excitaton energy using dipalmitoylphosphatidylethanolamine labelled with the donor N-(7-nitro-2,1,3-benzoxadiazol-4-yl) or the acceptor tetramethylrhodamine at the free amino group. The two methods yielded significantly different results. While recombination of contents could not be detected at Ca2+ concentrations below 2.5 mM the threshold concentration for lipid mixing was 1 mM. For saturating Ca2+ concentrations (>5 mM Ca2+) initial rates were higher by almost an order of magnitude for lipid mixing than for recombination of liposome contents. These observations indicate that the observation of rapid lipid mixing phenomena does not allow one to draw conclusions as to the fate of the enclosed volumes.  相似文献   

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