首页 | 本学科首页   官方微博 | 高级检索  
相似文献
 共查询到20条相似文献,搜索用时 31 毫秒
1.
The interaction of quinacrine with calf thymus DNA was monitored at several different ionic strengths using spectrophotometric and equilibrium dialysis techniques. The binding results can be explained, assuming each base pair is a potential binding site, using a model containing two negative cooperative effects: (1) ligand exclusion at binding sites adjacent to a filled binding site and (2) ligand–ligand negative cooperativity at adjacent filled binding sites. The logarithm of the observed equilibrium constant (Kobs) determined by this model varies linearily with log[Na+], as predicted by the ion condensation theory for polyelectrolytes. When the log Kobs plot is correlated for sodium release by DNA in the intercalation conformational change, the predicted number of ion pairs between the ligand and DNA is approximately two, as expected for the quinacrine dication. Even though Kobs depends strongly on ionic strength, the ligand negative cooperativity parameter ω was found to be indpendent of ionic strength within experimental error. This finding is also in agreement with the ion condensation theory, which predicts a relatively constant amount of condensed counterion on the DNA double helix over this ionic strength range. Drugs would, therefore, experience a relatively constant ionic environment when complexed to DNA even though the ionic conditions of the solvent could change considerably.  相似文献   

2.
The surface charge density (σ′0) and the binding constant of Ca++ with charged groups on the outer surface of the membrane (KCa) were calculated from experimentally determined values of the shift of the current-voltage characteristic curves of calcium currents in the membrane of rat spinal ganglion neurons: σ′0 = 0.15 ± 0.05 e/nm2 and KCa = 70 ± 10 liters/mole. Using a three-barrier model the energy profile of the calcium channel of the membrane of these neurons was calculated for Ca++, Ba++, Cd++, Mn++, Co++, and verapamil. The calcium current was shown to be determined mainly by the depth of the potential hole corresponding to the outer binding site of the calcium channel. It is concluded from the results that the outer binding site of the calcium channel contains only one carboxyl group.  相似文献   

3.
Proflavine binding may be used as a probe of the environment and interactions of DNA. In this paper we report the effects of the divalent cations Mg++ and putrescine and the trivalent cation spermidine on the proflavine–Na DNA binding equilibrium. Difference spectroscopy at 430 nm was used to determine apparent proflavine–DNA binding constants K at several concentrations of each cation for temperatures between 15 and 43°C, and at a constant total ionic strength of 0.1M. Mg++, putrescine, and spermidine all have greater effects on K than expected on the basis of ionic strength alone in the order spermidine > Mg++ ? putrescine. van't Hoff analysis of K(T) enabled calculation of ΔH° and ΔS°, which are affected differently by each cation. These differences are discussed qualitatively in terms of such concepts as release of condensed counterions, localized or unlocalized condensation, hydration, and restriction of molecular and internal rotation.  相似文献   

4.
Quercetin, a ubiquitous flavanoid, has numerous pharmacological effects, such as antioxidant and antitumor. Previous studies showed nucleic acids were the potential biological targets for antitumor medicine. For exploring the mechanism of DNA‐target medicine, the interaction between quercetin and calf thymus DNA was studied based on the method of spectrometry and simulation in our study. Firstly, the interaction between quercetin and calf thymus DNA was confirmed by fluorescence spectrometry. Furthermore, circular dichroism, fluorescence polarization, competitive displacement assay, and salt concentration dependence assay were applied to search the interaction mode of quercetin‐calf thymus DNA, which proved the existence of groove binding and electrostatic interaction. Meanwhile, quenching constant Ksv, binding constant Ka and the number of binding sites n was calculated, inferring that the fluorescence quenching occurred by static quenching process, and the main acting force was hydrogen bond. Finally, molecular docking was used to simulate and analyze the interaction between quercetin and calf thymus DNA.  相似文献   

5.
H Krakauer 《Biopolymers》1971,10(12):2459-2490
The binding of Mg ++ to polyadenylate (poly A), Polyuridylate(poly U), and their complexes, poly (A + U) and poly (A + 2U), was studied by means of a technique in which the dye eriochrome black T is used to measure the concentration of free Mg?. The apparent binding constant KX = [MgN]/[Mg++][N], N = site for Mg++ binding (the phosphate group of the nucleotide), was found to decrease rapidly as the extent of binding increased and, at low extents of binding, as the concentration of Na? increased in poly A, poly (A + U), and poly (A + 2U), and somewhat less so in poly U. Kx is generally in the range 104 > KX > 102. The cause of these dependences is apparently, primarily, the displacement of Na+ by Mg++ in poly U and poly (A + U) on the basis of the similarity of extents of displacement measured in this work and those measured potentiometrically. was calculated and was found to approach zero as the concentration of Na+ increased. In poly U, poly (A + U), and poly (A + 2U) at low ΔH′ v.H. > 0, about + 2 kcal/“mole.” In poly A, also at low salt, ΔH′ v.H. ≈ ?4 kcal/“mol” for the initial binding of Mg++, and increases to +2 kcal/“mol” at saturation. This enthalpic variation probably accounts for the anticooperativity in the binding of Mg++ not ascribable to the displacement of Na++.  相似文献   

6.
Interaction of magnesium ions with poly A and poly U   总被引:2,自引:0,他引:2  
The binding of Mg++ to poly A and poly U has been measured quantitatively by using the metallochromic indicator calmagite. The method is described in detail. It is shown that there is electrostatic interaction between the binding sites, viz., the phosphate groups, and the intrinsic association constant, for the specific binding can be determined. After extrapolation to zero ionic strength we find that, for the binding of Mg++ to poly A, kint = 4 × 104 and for that, to poly U, kint = 3 × 104. The intrinsic enthalpy of association is negative. The effect of Mg++ on the secondary structure of poly A and poly U has been studied by measuring the ultraviolet absorbance, optical rotatory dispersion and viscosity as a function of the amount of added Mg++ ions. It was found that Mg++ promotes the formation of a more ordered secondary structure by neutralizing or screening the negative charges. It is concluded from the absorbance measurements that for poly A at pH ? 7 and for poly U at pH >xs 9 this ordering involves stacking of the bases. Likewise, in solutions of UDP with a pH around 10, base stacking occurs on addition of Mg++.  相似文献   

7.
Binding constants and binding site sizes for the interactions of the polyamines spermine (+4), spermidine (+3), and putrecine (+2) with helical DNA have been determined as a function of ionic conditions and temperature by equilibrium dialysis using 14C-labeled polyamines. In addition, competition equilibrium dialysis has been used to determine binding parameters for the divalent cations putrescine and Mg2+ from the competitive effect of these ions on the binding of spermine or spermidine. In all cases, the logarithm of the binding constant (log Kobs) varies linearly with the logarithm of the monovalent salt concentration; the slopes d log Kobs/d log[NaCl] are proportional to the valence of the ligand, and values of the extrapolated binding constants at 1M NaCl obtained from the intercepts are small (of order 1–10M?1). In those cases examined, Kobs is insensitive to temperature; the free energy of binding is predominantly entropic. Consequently, polymines as DNA-binding ligands behave analogously to the oligolysìnes investigated previously [cf. Record, Lohman & de Haseth (1976) J. Mol. Biol. 107 , 145–158; Lohman, de Haseth & Record (1980) Biochemistry 19 , 3522–3530]. The interactions of these oligocations with DNA are predominantly electrostatic and are driven by the release of thermodynamically bound electrolyte ions from the vicinity of the DNA. The extent to which these oligocations are localized at individual phosphate binding sites or delocalized on the DNA molecule is currently not known.  相似文献   

8.
Abstract

The interaction between Tb(IV)-NR complex and herring sperm DNA in buffer solution of Tris-HCl was investigated with the use of acridine orange(AO) as a spectral probe. The binding modes and other information were provided by the UV–spectrophotometry and fluorescence spectroscopy. The thermodynamic functions expressed that the binding constants of Tb(IV)-NR complex with DNA was Kθ298.15K = 4.03?×?105?L·mol?1, Kθ310.15K =1.30?×?107?L·mol?1, and the ΔrGθ m 298.15?K=?3.20?×?104 J·mol?1. The scatchard equation suggested that the interaction mode between Tb(IV)-NR complex and herring sperm DNA is electrostatic and weak intercalation bindings. FTIR spectroscopy results also indicate that there is a specific interaction between the Tb(IV)-NR complex and the A and G bases of DNA.  相似文献   

9.
Ultraviolet-visible (UV-vis) spectra, fluorescence spectra, electrochemistry, and the thermodynamic method were used to discuss the interaction mode between the inclusion complex of hematoxylin with β-cyclodextrin and herring sperm DNA. On the condition of physiological pH, the result showed that hematoxylin and β-cyclodextrin formed an inclusion complex with binding ratio nhematoxylin:nβ-cyclodextrin = 1:1. The interaction mode between β-cyclodextrin-hematoxylin and DNA was a mixed binding, which contained intercalation and electrostatic mode. The binding ratio between β-cyclodextrin-hematoxylin and DNA was nβ-cyclodextrin -hematoxylin:nDNA = 2:1, binding constant was K? 298.15K = 5.29 × 104 L·mol?1, and entropy worked as driven force in this action.  相似文献   

10.
To evaluate the biological preference of [Yb(phen)2(OH2)Cl3](H2O)2 (phen is 1,10-phenanthroline) for DNA, interaction of Yb(III) complex with DNA in Tris–HCl buffer is studied by various biophysical and spectroscopic techniques which reveal that the complex binds to DNA. The results of fluorescence titration reveal that [Yb(phen)2(OH2)Cl3](H2O)2 has strongly quenched in the presence of DNA. The binding site number n, apparent binding constant K b, and the Stern–Volmer quenching constant K SV are determined. ΔH 0, ΔS 0, and ΔG 0 are obtained based on the quenching constants and thermodynamic theory (ΔH 0?>?0, ΔS 0?>?0, and ΔG 0?<?0). The experimental results show that the Yb(III) complex binds to DNA by non-intercalative mode. Groove binding is the preferred mode of interaction for [Yb(phen)2(OH2)Cl3](H2O)2 to DNA. The DNA cleavage results show that in the absence of any reducing agent, Yb(III) complex can cleave DNA. The antimicrobial screening tests are also recorded and give good results in the presence of Yb(III) complex.  相似文献   

11.
The 8-kDa subunit c of theE. coli F0 ATP-synthase proton channel was tested for Ca++ binding activity using a45Ca++ ligand blot assay after transferring the protein from SDS-PAGE gels onto polyvinyl difluoride membranes. The purified subunit c binds45Ca++ strongly with Ca++ binding properties very similar to those of the 8-kDa CF0 subunit III of choloroplast thylakoid membranes. The N-terminal f-Met carbonyl group seems necessary for Ca++ binding capacity, shown by loss of Ca++ binding following removal of the formyl group by mild acid treatment. The dicyclohexylcarbodiimide-reactive Asp-61 is not involved in the Ca++ binding, shown by Ca++ binding being retained in twoE. coli mutants, Asp61Asn and Asp61Gly. The Ca++ binding is pH dependent in both theE. coli and thylakoid 8-kDa proteins, being absent at pH 5.0 and rising to a maximum near pH 9.0. A treatment predicted to increase the Ca++ binding affinity to its F0 binding site (chlorpromazine photoaffinity attachment) caused an inhibition of ATP formation driven by a base-to-acid pH jump in whole cells. Inhibition was not observed when the Ca++ chelator EGTA was present with the cells during the chlorpromazine photoaffinity treatment. An apparent Ca++ binding constant on the site responsible for the UV plus chlorpromazine effect of near 80–100 nM was obtained using an EGTA-Ca++ buffer system to control free Ca++ concentration during the UV plus chlorpromazine treatment. The data are consistent with the notion that Ca++ bound to the periplasimic side of theE. coli F0 proton channel can block H+ entry into the channel. A similar effect occurs in thylakoid membranes, but the Ca++ binding site is on the lumen side of the thylakoid, where Ca++ binding can modulate acid-base jump ATP formation. The Ca++ binding to the F0 and CF0 complexes is consistent with a pH-dependent gating mechanism for control of H+ ion flux across the opening of the H+ channel.This work was supported in part by grants from the Department of Energy and the U.S. Department of Agriculture.On leave from the Institute of Soil Science and Photosynthesis, Russian Academy of Science, Pushchino, Russia.  相似文献   

12.
Unspecific binding of a protamine, namely fluorescein-labelled clupeine Z, to double-stranded calf thymus DNA was studied using fluorescence titration methods and chemical relaxation techniques. Both equilibrium and kinetic data have been analysed using general theoretical approaches discussed in the accompanying paper. The results agree well with the predictions made on the basis of a standard co-operative binding model.Basic parameters evaluated are the co-operative binding constant (K), the coefficient measuring co-operative interaction between nearest neighbours (q), the number of nucleotides occupied by one protamine molecule (n) and the rate constant of dissociation at the ends of bound ligand sequences (KD). Values obtained at 20 °C, pH 7.5 and 0.4 m-NaCl were K = 5.8 × 107m?1, q = 1700, n = 20 and KD = 0.29 s?1. They have been found to be sensitive to the concentration of added salt (NaCl). This effect apparently reflects the essentially electrostatic nature of the binding process. The results can be satisfactorily described in terms of competitive binding of sodium ions.  相似文献   

13.
We have isolated the MAP/tau proteins from twice-cycled chick brain microtubule preparations and demonstrated that they are responsible for the nitrocellulose DNA binding activity we and others have measured. Using the isolated MAP/tau proteins we then measured the apparent affinity constant Kapp for the homologous chick DNA interaction and found evidence for two equilibrium affinity classes-a Kapp = 6 × 107 M–1, responsible for the bulk of the DNA binding activity and a small (< 10%) higher affinity Kapp = 108 – 109 M–1, likely due to sequence specific binding protein species. Using the same chick brain MAP-tau protein, a heterologous interaction with D. melanogaster DNA, was found to possess just the lower affinity class-Kapp = 2 × 107 M–1. Under stringent binding conditions we carried out equilibrium nitrocellulose filter binding experiments in a ternary reaction mixture at constant MAP/tau protein and 35S radiolabelled chick DNA concentration using increasing and excess concentrations of competitor DNAs of different sources. The order of competitor strengths found was-chick DNA > mouse DNA > D. melanogaster = E. coli. DNA. These data and specifically the homologous DNA: protein case being the strongest competitor corroborate our previous studies using total microtubule protein and provide new evidence for a conserved interaction of a small DNA sequence class with MAP/tau protein species. Moreover, these data allow us to conclude that the conserved DNA sequence: MAP/tau protein interactions do not critically depend upon any energetic feature co-involving tubulin for their properties since tubulin is absent from these preparations.  相似文献   

14.
Summary The addition of agents that uncouple electron transfer from energy conservation (uncouplers) to state 4 mitochondria causes the following ion movements: K+ is extruded from the mitochondria in association with phosphate and possibly other anions, but not H+. Endogenous Ca++ is extruded from the mitochondria, and H+ moves in to counter-balance the Ca++ movement; some phosphate movement may be associated with Ca++ extrusion. The rate and extent of K+ extrusion induced by uncoupler is dependent on the concentrations of external phosphate and divalent ions. Phosphate induces K+ extrusion, while Mg++ and Mn++ inhibit it. TheV max of K+ transport is 300 moles K+/g protein per min. The K m for FCCP-induced potassium extrusion is 0.25 M at pH 7.4. The inhibitory effect of Mg++ is noncompetitive with respect to uncoupler concentration but competitive with respect to phosphate concentration. The experimental evidence does not support the existence of high H+ permeability in the presence of uncoupler. A correlation is observed between the rate of K+ extrusion and the energy reserves supplied from the high energy intermediate. The action of uncoupler in inducing K+ permeability is considered to arise through its action in depleting the energy reserves of mitochondria rather than through a specific activating effect of permeability by the uncoupler itself. The relationship of membrane potential to regulation of K+ permeability is discussed.  相似文献   

15.
In both photosynthetic (Pyrodinium bahamense, Gonyaulax polyedra, Pyrocystis Iunula, P. noctiluca, P. fusiformis) and nonphotosynthetic (Noctiluca miliaris) bioluminescent dinoflagellates chemical stimulation can by-pass mechanical stimulation. The effective ions are Ca++, K+, NH4+ and H+. Other chemicals found effective are those implicated in Ca++ transport or binding. There are interspecies differences in degrees of mechanical and chemical stimulability. Photoinhibition of mechanical stimulability is the result of two effects, the first a reduction in total bioluminescence potential and the second a decrease in mechanical stimulability resulting experimentally in a decreased rate of light emission. This latter effect can be reversed with Ca++ ions. Chemicals which bind Ca++ or displace Ca++ can mimic the effects of photoinhibition. The chemical inhibition of mechanical stimulability is also reversed by Ca++ ions. A scheme is proposed which is consistent for all species examined.  相似文献   

16.
Methods including spectroscopy, electronic chemistry and thermodynamics were used to study the inclusion effect between γ-cyclodextrin (CD) and vitamin K3(K3), as well as the interaction mode between herring-sperm DNA (hsDNA) and γ-CD-K3 inclusion complex. The results from ultraviolet spectroscopic method indicated that VK3 and γ-CD formed 1:1 inclusion complex, with the inclusion constant Kf = 1.02 × 104 L/mol, which is based on Benesi–Hildebrand's viewpoint. The outcomes from the probe method and Scatchard methods suggested that the interaction mode between γ-CD-K3 and DNA was a mixture mode, which included intercalation and electrostatic binding effects. The binding constants were K θ25°C = 2.16 × 104 L/mol, and Kθ37°C = 1.06 × 104 L/mol. The thermodynamic functions of the interaction between γ-CD-K3 and DNA were ΔrHmθ = ?2.74 × 104 J/mol, ΔrSmθ = 174.74 J·mol?1K?1, therefore, both ΔrHmθ (enthalpy) and ΔrSmθ (entropy) worked as driven forces in this action.  相似文献   

17.
Summary The Ca++-mediated increase in K+-permeability of intact red blood cells (Gardos effect) was initiated by exposing cells to known concentrations of Ca++ (using EGTA buffers) in the presence of the ionophore A23187. The potency of quinine, an inhibitor of the response, was found to depend on the external K+ concentration. In K+-free solutions the concentration of quinine to achieve 50% inhibition (K 50) was 5 m, but at 5mm K+ the required concentration was increased 20-fold to 100 m. An increase in internal Na+ had the opposite effect, allowing a high potency of quinine despite the presence of external K+. Alterations in the internal K+ level, on the other hand, were without effect on theK 50, suggesting that the membrane potential is not a factor. This conclusion is supported by the lack of effect on quinine inhibition of substitution of Cl by NO 3 , a considerably more permeant anion. The data are consistent with the hypothesis that quinine inhibits by competitively displacing K+ from an external binding site, the reported K+-activation site for the Ca++-mediated K+-permeability.  相似文献   

18.
The toxic interaction of melamine with herring sperm DNA (hs‐DNA) was investigated by using fluorescence and UV–vis absorption spectra techniques. The experimental results showed that the toxic interaction between melamine and hs‐DNA occurred. Fluorescence quenching experiments indicated the existence of electrostatic binding between melamine and hs‐DNA. The binding constants KA and the binding site numbers were calculated by means of the Stern–Volmer equation and were 9.8 × 104 L mol?1 and 1.3, respectively. Both the results of fluorescence spectra and UV–vis absorption spectra verified that there are electrostatic binding between melamine and hs‐DNA. The possibility in the presence of a classical intercalation binding mode could be ruled out by using DNA unwinding experiments. © 2010 Wiley Periodicals, Inc. J Biochem Mol Toxicol 24:323–329, 2010; View this article online at wileyonlinelibrary.com . DOI 10.1002/jbt.20341  相似文献   

19.
The binding of cupric ion (Cu++) to DNA was followed by spectrophotometry, melting profiles, and hydrodynamic techniques, in 0. 1M NaClO4 and at pH 5. 6. A small amount of Cu++ is bound specifically to bases (about 1 Cu++ per 20 nucleotides), in agreement with polarographic and EPR data. A preferential stabilization of G–C pairs and only a slight increase of the flexibility of the molecule were observed. In 5 × 10?3M NaClO4, a higher number of nonhomogeneous binding sites is found by spectrophotometry. It is concluded that at least two types of sites are available for Cu++. The first one, where Cu++ is chelating N7 of purines to phosphate, is observed only at low ionic strength and destabilizes the double helix. The second exists mainly at 0, 1M or higher ionic strength. All the sites are identical and could be attributed to two successive guanine residues in the same strand. Similar behavior was found for other divalent cations, e. g., Fe++, Mn++, and Co++.  相似文献   

20.
设为首页 | 免责声明 | 关于勤云 | 加入收藏

Copyright©北京勤云科技发展有限公司  京ICP备09084417号