共查询到20条相似文献,搜索用时 31 毫秒
1.
P Mohanakrishnan C F Chignell 《Comp. Biochem. Physiol. C, Comp. Pharmacol. Toxicol.》1984,79(2):321-323
The binding of copper and nickel to canine serum albumin has been studied using circular dichroism. In the 320-700 nm region, only a single positive extremum was observed at about 664-667.5 nm for copper bound to canine serum albumin. The intensity of this extremum was found to increase until a Cu2+/albumin molar ratio of 3 was reached. Further addition of Cu2+ led to a decrease in ellipticity. The absence of any extrema in the 560-570 and 480-510 nm regions showed that histidines were not involved in copper binding to canine albumin. In the case of nickel, initial binding was found to take place at the N-terminal tripeptide. At higher nickel to albumin molar ratios, circular dichroism spectra indicated the presence sulfur containing ligands but showed no evidence for the involvement of histidines. Canine serum albumin was found to bind six or more Cu2+ and Ni2+ ions with affinities that are lower than for human or bovine serum albumin. 相似文献
2.
The interaction of hemopexin with bilirubin was characterized by spectrophotometric, fluorimetric and circular dichroic techniques. Hemopexin rapidly forms an equimolar complex with libirubin that has an apparent dissociation constant Kd, of 7.5.10(-7) M. The association alters the absorption band of bilirubin near 150 nm, quenches the fluorescence of tryptophan residues of hemopexin, enhances the fluorescence of bilirubin, and induces strong ellipticity extrema in bilirubin of --60 . 10(3) deg . cm2 . dmol-1 at 465 nm and +70 . 10(3) deg . cm2 . dmol-1 at 415 nm. However, the conformation-sensitive ellipticity aband at 231 nm of hemopexin is not altered. In displacement experiments using circular dichroism, heme readily replaced bound bilirubin, indicating that bilirubin and heme are bound at the same site on hemopexin. Even at molar ratios of hemopexin to albumin of 3 to 1, human serum albumin removes bilirubin from hemopexin. Hemopexin is thus unlikely to have a role in the transport of bilirubin in serum. 相似文献
3.
The interaction between a thyroid hormone metabolite, 3-monoiodo-L-thyronine (3-T1) and bovine serum albumin (BSA) was investigated by using the CD method. An enhanced CD band was observed at the absorption wavelength region of 3-T1 around 293 nm suggesting the binding of 3-T1 to the BSA molecule. The ellipticity at 293 nm was measured at various molar ratios of 3-T1 to BSA, and the apparent binding constant and the maximum number of binding sites could be estimated as Kapp = 8.85 +/- 1.07 X 10(4) M-1 and n = 23.8 +/- 0.9 respectively. The CD of a mixture of BSA, 3-T1 and thyroxine (T4) was also studied at various pH's. The pH profile of the two characteristic CD bands at 293 nm and 320 nm, attributed to bound 3-T1 and T4, suggested that the optimum binding condition of 3-T1 was attained at alkaline pH of around 9, while that of T4 was attained over a wide pH range between 5-10. A significant role of the ionized 4'-hydroxyl group of 3-T1 in the binding reaction with BSA is also suggested. 相似文献
4.
pH-dependent bilayer destabilization by an amphipathic peptide 总被引:7,自引:0,他引:7
A 30-residue amphipathic peptide was designed to interact with uncharged bilayers in a pH-dependent fashion. This was achieved by a pH-induced random coil-alpha-helical transition, exposing a hydrophobic face in the peptide. The repeat unit of the peptide, glutamic acid-alanine-leucine-alanine (GALA), positioned glutamic acid residues on the same face of the helix, and at pH 7.5, charge repulsion between aligned Glu destabilized the helix. A tryptophan was included at the N-terminal as a fluorescence probe. The rate and extent of peptide-induced leakage of contents from large, unilamellar vesicles composed of egg phosphatidylcholine were dependent on pH. At pH 5.0 with a lipid/peptide mole ratio of 500/1, 100% leakage of vesicle contents occurred within 1 min. However, no leakage of vesicle contents occurred at pH 7.5. Circular dichroism measurements indicated that the molar ellipticity at 222 nm changed from about -4000 deg cm2 dmol-1 at pH 7.6 to -11,500 deg cm2 dmol-1 at pH 5.1, indicating a substantial increase in helical content as the pH was reduced. Changes in molar ellipticity were most significant over the same pH range where a maximum change in the extent and rate of leakage occurred. The tryptophan fluorescence emission spectra and the circular dichroism spectra of the peptide, in the presence of lipid, suggest that GALA did not associate with the bilayer at neutral pH. A change in the circular dichroism spectrum and a blue shift of the maximum of the tryptophan fluorescence emission spectra at pH 5.0, in the presence of lipid, indicated an association of GALA with the bilayer.(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献
5.
Effect of amino acid ion pairs on peptide helicity 总被引:12,自引:0,他引:12
The three ER ion pairs in the peptide acetyl-W(EAAAR)3A-amide were replaced in turn with the ion pairs EK, EO, DR, DK, and DO, where O represents an ornithine residue. The far-ultraviolet circular dichroic spectra of the six peptides measured in 10 mM NaCl at pH 2 and 0 degrees C form a nested set having an isodichroic point at 203 nm of -17,000 deg cm2 dmol-1. The ellipticity values of the six peptides at 222 nm range from -31,600 to -7400 deg cm2 dmol-1 in the order listed. Changing the pH of each peptide solution from 2 to 13 also generates a nested set of dichroic spectra with the same isodichroic values. Increasing the pH from 2 to 7 differentially increases the ellipticity at 222 nm in a single transition having an apparent pK of 4.1 for the E-containing peptides are 3.6 for the D-containing peptides. Increasing the pH beyond neutrality differentially decreases the ellipticity at 222 nm in a single transition having an apparent pK of greater than or equal to 13.2 for the R-containing peptides, 11.1 for the K-containing peptides, and 10.7 for the O-containing peptides. It is proposed that the difference in the ellipticity of the six peptides chiefly reflects the helix preferences for the variable residues supplemented by intrahelical electrostatic interactions in the neutral pH range. 相似文献
6.
Humra Athar Nisar Ahmad Saad Tayyab Mohammad A. Qasim 《International journal of biological macromolecules》1999,25(4):1547-358
Bilirubin–albumin solution gave an emission spectrum in the wavelength range 500–600 nm with emission maxima at 528 nm when excited at 487 nm. The magnitude of fluorescence intensity increased on increasing bilirubin/albumin molar ratio. At three different albumin concentrations, namely, 1.0, 2.5 and 10.0 μM, there was an initial linear increase in fluorescence up to a molar ratio 1.0 in all cases beyond which it sloped off or decreased. This fluorescence enhancement was used to calculate the binding parameters of bilirubin–albumin interaction and the value of binding constant was found to be 1.72×107 l/mol similar to the published values obtained with other methods. Different serum albumins, namely, human (HSA), goat (GSA), pig (PSA) and dog serum albumins (DSA) bound bilirubin with almost the same affinity when studied by the technique of fluorescence enhancement. Bilirubin–albumin interaction was also studied at different pH and ionic strengths. There was a decrease in bilirubin–albumin complex formation on either decreasing the pH from 9.0 to 7.0 or increasing the ionic strength from 0.15 to 1.0. These results suggest that the technique of fluorescence enhancement can be used successfully to study the bilirubin–albumin interaction. 相似文献
7.
J Magdalou C Thirion M Balland G Siest 《The International journal of biochemistry》1985,17(10):1103-1107
Ultraviolet circular dichroism spectrum of purified NADPH cytochrome P-450 reductase was characterized by two negative bands centered at 208 and 222 nm. The approximation of the alpha-helical content from the value of the mean residue ellipticity at 222 nm indicated 28% of alpha-helical structures. Heat inactivation of the enzyme was associated to a drastic change in the secondary structure of the protein. Membrane reconstitution experiments by inclusion of the enzyme into liposomes revealed that the conformation of NADPH cytochrome P-450 reductase was sensitive to its phospholipid environment. Egg lecithin as well as synthetic phosphatidylcholines, at the optimal phospholipid-enzyme molar ratio 200, was able to increase up to 37% the mean residue ellipticity at 222 nm. Addition of phosphatidylserine or phosphatidylethanolamine produced no effect. Non-ionic detergent such as Emulgen 913 weakly enhanced the mean residue ellipticity. 相似文献
8.
Optical properties of hyaluronic acid. Ultraviolet circular dichroism and optical rotatory dispersion 总被引:3,自引:0,他引:3
The molar optical rotation at 220 nm and ellipticity values at 210 nm of both sodium hyaluronate and hyaluronic acid are greatly enhanced in comparison to the values for the monomeric units and oligosaccharides indicating a degree of preferred order. With increasing hydrogen ion concentration, there is no appreciable change in the 210 nm circular dichroic band, but the second circular dichroic band below pH 4 changes abruptly to the positive side and reaches a maximum value at pH 2·5. This positive circular dichroic band of hyaluronic acid is temperature and concentration dependent. The major change in sign and position of the second circular dichroic band of hyaluronic acid below pH 4 is attributed to the conformational change of a single polysaccharide chain or to a chain-chain interaction. The results indicate that increase in concentration or decrease in temperature and in the ionization of carboxyl group promotes the formation of ordered cross-link regions. The conformational changes found in solution have been interpreted as an order-disorder transition in the crosslink regions based on the interconversion of random coil and double helix. 相似文献
9.
Abstract— A highly purified basic protein prepared from pig brain was studied by circular dichroism and viscometry. The circular dichroism spectrum of the protein in 50% (v/v) aqueous n-propanol showed two negative bands (at 217-220 nm and 204 nm); the spectrum in 90% (v/v) aqueous trifluoroethanol similarly showed two negative bands (at 217 nm and at 206 nm). The molar ellipticity of this protein in aqueous solvents was relatively flat and no negative bands were observed above 200 nm. The ηap C of the protein from pig brain was 0-12 as C → O in tris buffer (0-1 M, pH 7-8). On the basis of these studies we concluded that the protein is asymmetric and extended in aqueous systems and contains a maximum of 20 per cent α-helix in trifluoroethanol. 相似文献
10.
Circular dichroism properties of SV40 virions, isolated minichromosomes from virions, and SV40 Form I (supercoiled) DNA were studied in a buffer of low ionic strength. The isolated minichromosomes are compact as judged by sedimentation and electron microscopy. The molar ellipticity at 284 nm of the virion, which may be regarded as a minichromosome in its native state, is about 1500 deg cm2/dmol phosphate; this value is in the same range as that reported for core particles (1300–2000) isolated from different sources. When the viral capsid is removed, there is a small increase in the molar ellipticity to about 2000. However, both of these values are much lower than that found for SV40 supercoiled DNA (about 8200). The results strongly suggest that the linker DNA of the native whole chromatin contributes in a similar fashion to the circular dichroic ellipticity as the core DNA. 相似文献
11.
Md. Khurshid Alam Khan Md. Hamidur Rahaman Md. Imtaiyaz Hassan Tej P. Singh Ali A. Moosavi-Movahedi Faizan Ahmad 《Journal of biological inorganic chemistry》2010,15(8):1319-1329
It has already been shown that the mutant Leu94Gly of horse cytochrome c exists in a molten globule (MG) state. We have carried out studies of reversible folding and unfolding induced by LiCl of
this mutant at pH 6.0 and 25 °C by observing changes in the difference molar absorption coefficient at 402 nm, the mean residue
ellipticity at 222 nm, and the difference mean residue ellipticity at 409 nm. This process is a three-state process when measured
by these probes. The stable folding intermediate state has been characterized by far- and near-UV circular dichroism, tryptophan
fluorescence, 8-anilino-1-naphthalenesulfonic acid binding, and dynamic light scattering measurements, which led us to conclude
that the intermediate is a premolten globule (PMG). Analysis of the reversible unfolding transition curves for the stability
of different states in terms of the Gibbs free energy change at pH 6.0 and 25 °C led us to conclude that the MG state is more
stable than the PMG state by 5.4 ± 0.1 kcal mol−1, whereas the PMG state is more stable than the denatured (D) state by only 1.1 ± 0.1 kcal mol−1. A comparison of the conformational and thermodynamic properties of the LiCl-induced PMG state at pH 6.0 with those of the
PMG state induced by NaCl at pH 2.0 suggests that a similar PMG state is obtained under both denaturing conditions. Differential
scanning calorimetry measurements suggest that heat induces a reversible two-state transition between MG and D states. 相似文献
12.
The circular dichroic spectrum of synaptosomal membranes was highly reproducible and qualitatively similar to that of erythrocyte membranes. The spectrum exhibited a minimum at 224 nm, a shoulder at 212 nm and a maximum at 195 nm. The mean molar ellipticity at the maximum and minimum was approximately +8000 and –8000 respectively. The protein components were the dominant source of the CD signal. Quantitative estimates showed negligible contributions to the spectrum from cholesterol, phosphatidyl serine and TV-acyl sugars. Phospholine iodide, eserine, decamethonium, tetramethyl ammonium chloride and acetylcholine at concentrations of 10-3 and 10-4M did not produce detectable perturbations of the membrane circular dichroism. The circular dichroic spectrum of d-tubocurarine exhibited a maximum at 198 nm and a minimum at 212 nm. Addition of d-tubocurarine to membrane suspensions in the cuvette yielded a complex spectrum representing the result of a simple additive combination of the circular dichroic spectra of d-tubocurarine and the membranes. However, preincubation of the membranes with 3 x 10-3m d-tubocurarine at 0-4°C for 5-10 min followed by sedimentation, several washes and resuspension resulted in a circular dichroic spectrum which appeared to involve no change in the membrane contribution, but there was a substantial decrease in the molar ellipticity of the d-tubocurarine remaining with the membranes. 相似文献
13.
The fluorescence emission spectrum of soybean dihydrofolate reductase suggests that the emitting tryptophan residues are situated in a hydrophobic microenvironment. The dissociation constants determined from fluorescence and circular dichroism data reveal that the soybean enzyme has a lower affinity for substrates and substrate analogs than that determined for dihydrofolate reductases isolated from other sources. The binding of methotrexate to the soybean enzyme does not affect the binding of NADPH. Similarly, the binding of NADPH has no effect on subsequent methotrexate binding. Polarimetric study indicates that the enzyme has a low (ca. 5%) α-helical content. Addition of dihydrofolate to the soybean enzyme results in the generation of a positive ellipticity band at 298 nm with a molar ellipticity, [θ], of 186,000, whereas the binding of folate induces a negative ellipticity band at 280 nm with [θ] of ?181,000. The qualitative and quantitative differences in the circular dichroism of the enzyme-dihydrofolate and enzyme-folate complexes indicate that the mode of binding of these ligands may be different. The formation of an enzyme-NADPH complex is accompanied by a negative Cotton effect at 270 nm. These studies indicate that the binding of substrates or inhibitors causes significant conformational changes in the enzyme and also leads to the formation of a number of spectroscopically identifiable complexes. 相似文献
14.
We studied the effect of various anions (of acids and salts) on the acid denatured state of HSA by near-UV circular dichroism (CD), far-UV CD, 1-anilinonaphthalene-8-sulfonate (ANS) binding, tryptophan fluorescence and thermal transition. Addition of different acids and salts caused an induction of alpha-helical structure as evident from the increase in the mean residue ellipticity (MRE) value at 222 nm and loss of ANS binding sites exhibited by the decrease in the ANS fluorescence intensity at 480 nm. However, the concentration range of acids/salts required to bring about the transition varied greatly among different acids and salts. Among various acids/salts tested, K(3)Fe(CN)(6) was found to be most effective whereas HCl and KCl were least effective in inducing the properties close to native structure. Further, they followed the electroselectivity series. The near-UV CD spectra showed an increase in MRE towards the native state, whereas the tryptophan fluorescence emission spectra produced a red shift of about 6 nm on addition of KClO(4). The temperature-induced transition in the presence of 40 mM KClO(4) monitored by ellipticity measurements at 222 nm was characterized by the presence of an intermediate state in the temperature range 30-50 degrees C having abundant secondary structure. These results suggest that human serum albumin at low pH and in the presence of acids or salts exists in a partially folded state characterized by native-like secondary structure and tertiary folds. 相似文献
15.
The effect of a cationic detergent, lauryl pyridiniumchloride (LPC), and an anionic one, sodium n-octylbenzenesulfonate (SOBS), on the conformation of unfractionated Escherichia coli tRNA was investigated at various molar ratios of detergent to tRNA (D/R) in the presence and absence of Mg2+ and Na+ ions by measuring the circular dichroism (CD) at 265 nm and 340 nm, which reflects conformational change involving base pairs and/or base stacking, and the disymmetry in the vicinity of 4-thiouridylate (4-TU), respectively. In the presence of Mg2+ and Na+ ions, the tRNA retains its native structure even in the presence of high molar ratios of detergent to tRNA (D/R congruent to 40 at 265 nm and D/R congruent to 20 at 340 nm). However, in the absence of these metal ions, the ellipticity at 340 nm was very sensitive to LPC concentration and decreased from 5,600 to nearly--1,600 at 25 degrees C with the increase of D/R ratios up to 20, and a similar decrease in the ellipticity at 340 nm was observed on thermal denaturation. This result suggests that the local environment involving the 4-TU region might be readily influenced by LPC, reflecting a large conformational change. However, no effect was observed in the case of the SOBS: tRNA system. On the other hand, secondary base pairing and/or base stacking structure was virtually invariant on adding both LPC and SOBS even at high D/R ratios in the absence of Mg2+ and Na+ ions. 相似文献
16.
The kinetics of regain of the native ellipticity in the far- and near-UV spectra have been investigated during the refolding at pH 7.8 and 20 degrees C of guanidine-unfolded, nonreduced hen egg white lysozyme. Stopped-flow studies showed that the ellipticities at 260 and 289.5 nm exhibit biphasic kinetics with rate constants of about 50 s-1 and 2.5 s-1 for the rapid and slow phase, respectively. The ellipticity in the far-UV obeyed triphasic kinetics. In addition to a rapid and a slow phase with rate constants similar to those observed in the near-UV, a "burst" of ellipticity was shown to occur in the dead time of the experiments. The effects of low pH and of concentrations of guanidine ranging from 0.075 to 1.5 M on the rapid and slow rate constants were studied. Under all conditions investigated, the rate constants observed in the far- and near-UV for a given phase were the same, thus suggesting that the molecular events observed in the two regions of the UV spectrum are either identical or strongly coupled. Continuous-flow experiments at different wavelengths between 214 and 240 nm under conditions where the dead time for the observation was only 4 ms, followed by a detailed analysis of the kinetics of ellipticity change at each wavelength, provided the spectrum of the molecular species formed at the end of the burst phase. This spectrum was found to closely fit that predicted from the secondary structure of native lysozyme.(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献
17.
H W Dirr 《Biochemistry international》1987,14(4):727-733
The influence of pH and long-chain fatty acids on the interaction between aflatoxin B1 and human albumin was investigated by fluorescence spectroscopy. Both the binding of aflatoxin B1 to albumin and the fluorescence of albumin-bound aflatoxin are pH-dependent over the pH range of 6-9.5. The data indicates that the carcinogen has a higher affinity for the basic(B) than for the neutral(N) conformation of human albumin. Palmitic, stearic and oleic acids up to a molar ratio of 2 over albumin, increases the binding strength of aflatoxin B1 by means of an allosteric mechanism. Furthermore, the pH-dependence of the aflatoxin-albumin interaction is affected by the presence of oleic acid by narrowing the pH range over which the dependence occurs. At molar ratios of oleic acid to albumin in excess of 4.25 at pH6, 3.1 at pH7.4 and 2.4 at pH9 cause a decrease in aflatoxin B1 fluorescence as a result of reduced binding to albumin. 相似文献
18.
Abstract The binding of the benzodioxolo-benzoquinolizine alkaloid, berberine chloride to natural and synthetic DNAs has been studied by intrinsic and extrinsic circular dichroic measurements. Binding of berberine causes changes in the circular dichroism spectrum of DNA as shown by the increase of molar ellipticity of the 270nm band, but with very little change of the 240nm band. The molar ellipticity at the saturation depends strongly on the base composition of DNA and also on salt concentration, but always larger for the AT rich DNA than the GC rich DNA The features in the circular dichroic spectral changes of berberine-synthetic DNA complexes were similar to that of native DNA but depends on the sequence of base pairs. On binding to DNA and polynucleotides, the alkaloid becomes optically active. The extrinsic circular dichroism developed in the visible absorption region (300–500nm) for the berberine-DNA complexes shows two broad spectral bands in the regions 425–440nm and 340–360nm with the maximum varying depending on base composition and sequence of DNA While the 425nm band shows less variation on the binding ratio, the 360nm band is remarkably dependent on the DNA/alkaloid ratio. The generation of the alkaloid associated extrinsic circular dichroic bands is not dependent on the base composition or sequence of base pairs, but the nature and magnitude of the bands are very much dependent on these two factors and also on the salt concentration. The interpretation of the results with respect to the modes of the alkaloid binding to DNA are presented. 相似文献
19.
The binding of the benzodioxolo-benzoquinolizine alkaloid, berberine chloride to natural and synthetic DNAs has been studied by intrinsic and extrinsic circular dichroic measurements. Binding of berberine causes changes in the circular dichroism spectrum of DNA as shown by the increase of molar ellipticity of the 270nm band, but with very little change of the 240nm band. The molar ellipticity at the saturation depends strongly on the base composition of DNA and also on salt concentration, but always larger for the AT rich DNA than the GC rich DNA. The features in the circular dichroic spectral changes of berberine-synthetic DNA complexes were similar to that of native DNA, but depends on the sequence of base pairs. On binding to DNA and polynucleotides, the alkaloid becomes optically active. The extrinsic circular dichroism developed in the visible absorption region (300-500nm) for the berberine-DNA complexes shows two broad spectral bands in the regions 425-440nm and 340-360nm with the maximum varying depending on base composition and sequence of DNA. While the 425nm band shows less variation on the binding ratio, the 360nm band is remarkably dependent on the DNA/alkaloid ratio. The generation of the alkaloid associated extrinsic circular dichroic bands is not dependent on the base composition or sequence of base pairs, but the nature and magnitude of the bands are very much dependent on these two factors and also on the salt concentration. The interpretation of the results with respect to the modes of the alkaloid binding to DNA are presented. 相似文献
20.
Moza B Qureshi SH Islam A Singh R Anjum F Moosavi-Movahedi AA Ahmad F 《Biochemistry》2006,45(14):4695-4702
We have carried out denaturation studies of bovine cytochrome c (cyt c) by LiClO4 at pH 6.0 and 25 degrees C by observing changes in difference molar absorbance at 400 nm (Deltaepsilon400), mean residue ellipticities at 222 nm ([theta]222) and difference mean residue ellipticity at 409 nm (Delta[theta]409). The denaturation is a three-step process when measured by Deltaepsilon400 and Delta[theta]409, and it is a two-step process when monitored by [theta]222. The stable folding intermediate state has been characterized by near- and far-UV circular dichroism, tryptophan fluorescence, 8-anilino-1-naphthalene sulfonic acid (ANS) binding, and intrinsic viscosity measurements. A comparison of the conformational and thermodynamic properties of the LiClO4-induced molten globule (MG) state with those induced by other solvent conditions (e.g., low pH, LiCl, and CaCl2) suggests that LiClO4 induces a unique MG state, i.e., (i) the core in the LiClO4-induced state retains less secondary and tertiary structure than that in the MG states obtained in other solvent conditions, and (ii) the thermodynamic stability associated with the LiClO4-induced process, native state <--> MG state, is the same as that observed for each transition between native and MG states induced by other solvent conditions. 相似文献