共查询到20条相似文献,搜索用时 31 毫秒
1.
S. Benini Marco Borsari S. Ciurli Alexander Dikiy M. Lamborghini 《Journal of biological inorganic chemistry》1998,3(4):371-382
Direct cyclic voltammetry and 1H NMR spectroscopy have been combined to investigate the electrochemical and spectroscopic properties of cytochrome c
553 isolated from the alkaliphilic soil bacterium Bacillus pasteurii. A quasi-reversible diffusion-controlled redox process is exhibited by cytochrome c
553 at a pyrolitic graphite edge microelectrode. The temperature dependence of the reduction potential, measured using a non-isothermal
electrochemical cell, revealed a discontinuity at 308 K. The thermodynamic parameters determined in the low-temperature range
(275–308 K;ΔS°′=–162.7±1.2 J mol–1 K–1, ΔH°′=–53.0±0.5 kJ mol–1, ΔG°′=–4.5±0.1 kJ mol–1, E°′=+47.0±0.6 mV) indicate the presence of large enthalpic and entropic effects, leading, respectively, to stabilization and
destabilization of the reduced form of cytochrome c
553. Both effects are more accentuated in the high-temperature range (308–323 K;ΔS°′=–294.1±8.4 J mol–1 K–1, ΔH°′=–93.4±3.1 kJ mol–1, ΔG°′=–5.8±0.6 kJ mol–1, E°′=+60.3±5.8 mV), with the net result being a slight increase of the standard reduction potential. These thermodynamic parameters
are interpreted using the compensation theory of hydration of biopolymers as indicating the extrusion, upon reduction, of
water molecules from the hydration sphere of the cytochrome. The low-T and high-T conformers differ by the number of water molecules in the solvation sphere: in the high-T conformer, the number of water molecules extruded upon reduction increases, as compared to the low-T conformer. The ionic strength dependence of the reduction potential at 298 K, treated within the frame of extended Debye-Hückel
theory, yields values of E
°′
(I=0)
=–25.4±1.4 mV, z
red=–11.3, and z
ox=–10.3. The pH dependence of the reduction potential at 298 K shows a plateau in the pH range 7–10 and an increase at more
acidic pH, allowing the calculation of pK
O=5.5 and pK
R=5.7, together with the estimate of the reduction potentials of completely protonated (+71 mV) and deprotonated (+58 mV) forms
of cytochrome c
553. 1H NMR spectra of the oxidized paramagnetic cytochrome c
553 indicate the presence of a His-Met axial coordination of the low-spin (S=1/2) heme iron, which is maintained in the temperature interval 288–340 K at pH 7 and in the pH range 4.8–10.0 at 298 K.
The temperature dependence of the hyperfine-shifted signals shows both Curie-type and anti-Curie-type behavior, with marked
deviations from linearity, interpreted as indicating the presence of a fast equilibrium between the low-T and high-T conformers, having slightly different heme electronic structures resulting from the T-induced conformational change. Increasing the NaCl concentration in the range 0–0.2 M causes a slight change of the 1H NMR chemical shifts of the hyperfine-shifted signals, with no influence on their linewidth. The calculated lower limit value
of the apparent affinity constant for specific ion binding is estimated as 5.2±1.1 M–1. The pH dependence of the isotropically shifted 1H NMR signals of the oxidized cytochrome displays at least one ionization step with pK
O=5.7. The thermodynamic and spectroscopic data indicate a large solvent-derived entropic effect as the main cause for the
observed low reduction potential of B. pasteurii cytochrome c
553.
Received: 9 January 1998 / Accepted: 8 April 1998 相似文献
2.
M.I. Rajoka Khalil-ur- Rehman Tehmina Tabish M.A. Zia 《World journal of microbiology & biotechnology》2006,22(3):289-291
Purified uricase from a caprine kidney, possessed K
m
and V
max values of 1.1 mg ml−1 and 3512 IU (mg protein)−1 for uric acid hydrolysis, respectively. The optimum temperature and pH for catalytic activity were 40 °C and 8.5, respectively.
The activation energy for formation of ES complex was 13.6 kJ mol−1. Enthalpy (ΔH*), entropy of activation (ΔS*) and Gibbs free energy demand of uricase inactivation were 62.8 kJ mol−1, −102 J mol−1 K−1 and 104.3 kJ mol−1, respectively. Gibbs free enrgy demand for substrate binding and transition state stabilization were also determined which
were comparable with those for themostable enzymes. 相似文献
3.
Hiroshi Takashima Ayako Araki Keiko Takemoto Naokazu Yoshikawa Keiichi Tsukahara 《Journal of biological inorganic chemistry》2006,11(3):316-324
In order to understand the detailed mechanism of the stereoselective photoinduced electron-transfer (ET) reactions of zinc-substituted
myoglobin (ZnMb) with optically active molecules by flash photolysis, we designed and prepared new optically active agents,
such as N,N′-dimethylcinchoninium diiodide ([MCN]I2) and N,N′-dimethylcinchonidinium diiodide ([MCD]I2). The photoexcited triplet state of ZnMb, 3(ZnMb)*, was successfully quenched by [MCN]2+ and [MCD]2+ ions to form the radical pair of ZnMb cation (ZnMb·+) and reduced [MCN]·+ and [MCD]·+, followed by a thermal back ET reaction to the ground state. The rate constants (k
q) for the ET quenching at 25 °C were obtained as k
q(MCN)=(1.9±0.1)×106 M−1 s−1 and k
q(MCD)=(3.0±0.2)×106 M−1 s−1, respectively. The ratio of k
q(MCD)/k
q(MCN)=1.6 indicates that the [MCD]2+ preferentially quenches 3(ZnMb)*. The second-order rate constants (k
b) for the thermal back ET reaction from [MCN]·+ and [MCD]·+ to ZnMb·+ at 25 °C were k
b(MCN)=(0.79±0.04)×108 M−1 s−1 and k
b(MCD)=(1.0±0.1)×108 M−1 s−1, respectively, and the selectivity was k
q(MCD)/k
q(MCN)=1.3. Both quenching and thermal back ET reactions are controlled by the ET step. In the quenching reaction, the energy
differences of ΔΔH
≠(MCD–MCN) and ΔΔS
≠(MCD–MCN) at 25 °C were obtained as −1.1 and 0 kJ mol−1, respectively. On the other hand, ΔΔH
≠(MCD–MCN)=11±2 kJ mol−1 and TΔΔS
≠(MCD–MCN)=−10±2 kJ mol−1 were given in the thermal back ET reaction. The highest stereoselectivity of 1.7 for [MCD]·+ found at low temperature (10 °C) was due to the ΔΔS
≠ value obtained in the thermal back ET reaction.
Electronic Supplementary Material Supplementary material is available for this article at and is accessible for authorized users. 相似文献
4.
Durmaz-Sam S Sayar NA Topal-Sarikaya A Sayar AA 《Bioprocess and biosystems engineering》2011,34(8):997-1005
The potential of the dried yeast, wild-type Schizosaccharomyces pombe, to remove Ni(II) ion was investigated in batch mode under varying experimental conditions including pH, temperature, initial
metal ion concentration and biosorbent dose. Optimum pH for biosorption was determined as 5.0. The highest equilibrium uptake
of Ni(II) on S. pombe, q
e, was obtained at 25 °C as 33.8 mg g−1. It decreased with increasing temperature within a range of 25–50 °C denoting an exothermic behaviour. Increasing initial
Ni(II) concentration up to 400 mg L−1 also elevated equilibrium uptake. No more adsorption took place beyond 400 mg L−1. Equilibrium data fitted better to Langmuir model rather than Freundlich model. Sips, Redlich–Peterson, and Kahn isotherm
equations modelled the investigated system with a performance not better than Langmuir. Kinetic model evaluations showed that
Ni(II) biosorption process followed the pseudo-second order rate model while rate constants decreased with increasing temperature.
Gibbs free energy changes (ΔG°) of the system at 25, 30, 35 and 50 °C were found as −1.47E + 4, −1.49E + 4, −1.51E + 4, and −1.58E + 4 J mol−1, respectively. Enthalpy change (ΔH°) was determined as −2.57E + 3 J mol−1 which also supports the observed exothermic behaviour of the biosorption process. Entropy change (ΔS°) had a positive value (40.75 J mol−1 K−1) indicating an increase in randomness during biosorption process. Consequently, S. pombe was found to be a potential low-cost agent for Ni(II) in slightly acidic aqueous medium. In parallel, it has been assumed
to act as a separating agent for Ni(II) recovery from its aqueous solution. 相似文献
5.
Studied was the effect of temperature in the range 12–46 °C on the rate of bacterial decolorization of the mono-azo dye Acid
Orange 7 by Alcaligenes faecalis 6132 and Rhodococcus erythropolis 24. With both strains the raise of temperature led to a corresponding raise of decolorization rate better manifested by R. erythropolis. The analysis of the Arrhenius plot revealed a break near the middle of the temperature range. The regression analysis showed
practically complete identity of the observed break point temperatures (T
BP): 20.7 °C for Alc. faecalis and 20.8 °C for R. erythropolis. The values of the activation energy of the decolorization reaction (E
a) were found to depend on both the organism and the temperature range. In the range below T
BP the estimated values of E
a were 138 ± 7 kJ mol−1 for Alc. faecalis and 160 ± 8 kJ mol−1 for R. erythropolis. In the range above T
BP they were 54.2 ± 1.8 kJ mol−1 for Alc. faecalis and 37.6 ± 4.1 kJ mol−1 for R. erythropolis. Discussed are the possible reasons for the observed abrupt change of the activation energy. 相似文献
6.
The interaction between benzophenone (BP) and bovine serum albumin (BSA) was investigated by the methods of fluorescence spectroscopy
combined with UV–Vis absorption and circular dichroism (CD) measurements under simulative physiological conditions. The experiment
results showed that the fluorescence quenching of BSA by BP was resulted from the formation of a BP–BSA complex and the corresponding
association constants (K
a) between BP and BSA at four different temperatures had been determined using the modified Stern–Volmer equation. The enthalpy
change (ΔH) and entropy change (ΔS) were calculated to be –43.73 kJ mol−1 and −53.05 J mol−1 K−1, respectively, which suggested that hydrogen bond and van der Waals force played major roles in stabilizing the BP–BSA complex.
Site marker competitive experiments indicated that the binding of BP to BSA primarily took place in site I (sub-domain IIA).
The conformational investigation showed that the presence of BP decreased the α-helical content of BSA and induced the slight unfolding of the polypeptides of protein, which confirmed some micro-environmental
and conformational changes of BSA molecules. 相似文献
7.
P. O. Vardevanyan A. P. Antonyan G. A. Manukyan A. T. Karapetyan A. K. Shchyolkina O. F. Borisova 《Molecular Biology》2000,34(2):272-276
Isotherms of the EtBr adsorption on native and denatured poly(dA)poly(dT) in the temperature interval 20–70°C were obtained.
The EtBr binding constants and the number of binding sites were determined. The thermodynamic parameters of the EtBr intercalation
complex upon changes of solution temperature 20–48°C were calculated: 1.0·106 M−1≤K≤1.4·106 M−1, free energy ΔG
o=−8.7±0.3 kcal/mol, enthalpy ΔH
o≅0, and entropy ΔS
o=28±0.5 cal/(mol deg). UV melting has shown that the melting temperature (T
m) of EtBr-poly(dA)poly(dT) complexes (μ=0.022,4.16·10−5 M EtBr) increased by 17°C as compared with the ΔT
m of free homopolymer, whereas the half-width of the transition (T
m) is not changed. It was shown for the first time that EtBr forms complexes of two types on single-stranded regions of poly(dA)poly(dT)
denatured at 70°C: strong (K
1=1.7·105 M−1; ΔG
o=−8.10±0.03 kcal/mol) and weak (K
2=2.9·103 M−1; ΔG
o=−6.0±0.3 kcal/mol).The ΔG
o of the strong and weak complexes was independent of the solution ionic strength, 0.0022≤μ≤0.022. A model of EtBr binding
with single-stranded regions of poly(dA)poly(dT) is discussed. 相似文献
8.
To elucidate determinants of thermostability and folding pathways of the intrinsically stable proteins from extremophilic
organisms, we are studying β-glucosidase from Pyrococcus furiosus. Using fluorescence and circular dichroism spectroscopy, we have characterized the thermostability of β-glucosidase at 90°C,
the lowest temperature where full unfolding is achieved with urea. The chemical denaturation profile reveals that this homotetrameric
protein unfolds at 90°C with an overall ΔG° of ∼ 20 kcal mol−1. The high temperatures needed to chemically denature P. furiosus β-glucosidase and the large ΔG° of unfolding at high temperatures shows this to be one of the most stable proteins yet characterized.
Unfolding proceeds via a three-state pathway that includes a stable intermediate species. Stability of the native and intermediate
forms is concentration dependent, and we have identified a dimeric assembly intermediate using high temperature native gel
electrophoresis. Based on this data, we have developed a model for the denaturation of β-glucosidase in which the tetramer
dissociates to partially folded dimers, followed by the coupled dissociation and denaturation of the dimers to unfolded monomers.
The extremely high stability is thus derived from a combination of oligomeric interactions and subunit folding. 相似文献
9.
Chlorogenic acid, 3’-O-caffeoyl D-quinic acid, is an inherent ligand present inHelianthus annuus L. The effect of pH on chlorogenic acid binding to helianthinin suggests that maximum binding occurs at pH 6.0. The protein-polyphenol
complex precipitates as a function of time. The association constant of the binding of chlorogenic acid to helianthinin, determined
by equilibrium dialysis, at 31°C has a value of 3.5 ± 0.1 × 104M−-1 resulting in a ΔG value of − 6.32 ± 0.12 kcal /mol. The association constantK
ais 1.0 ± 0.1 × 104M−1 as determined by ultraviolet difference spectral titration at 25°C with ΔG° of -5.46 ± 0.06 kcal/mol. From fluorescence spectral
titration at 28°C, theK
avalue is 1.38 ± 0.1 × 1 0 4M−1 resulting in a ΔG of − 5.70 ± 0.05 kcal/mol. The total number of binding sites on the protein are 420 ± 50 as calculated
from equilibrium dialysis. Microcalorimetric data of the ligand-protein interaction at 23°C suggests mainly two classes of
binding. The thermal denaturation temperature,T
mof the protein decreases from 76°C to 72°C at 1 × 10−3M chlorogenic acid concentration upon complexation. This suggests that the complexation destabilizes the protein. The effect
of temperature onK
aof chlorogenic acid shows a nonlinear increase from 10.2°C to 45°C. Chemical modification of both lysyl and tryptophanyl residues
of the protein decreases the strength of binding of chlorogenic acid. Lysine, tryptophan and tyrosine of protein are shown
to be present at the binding site. Based on the above data, it is suggested that charge-transfer complexation and entropically
driven hydrophobic interaction are the predominant forces that are responsible for binding of chlorogenic acid to the multisubunit
protein, helianthinin.
Publication No. 324. 相似文献
10.
The salt dependence of the binding constant (K) and enthalpic (ΔH) and entropic (ΔS) components for magnesium binding to
poly-RNA was determined as a function of the concentration and identity of monovalent counter ions (M+). Both ΔH and ΔS were found to vary linearly with ln [M+]. A theoretical analysis of the experimental data revealed that the temperature dependence of the product of the density
of bound counter ions and the electrostatic interaction parameter, δ(m′ψ)/δT, is non-negligible, although it has previously
been ignored. The sign of δ(m′ψ)/δT was negative for poly(A) and positive for poly(U), indicating that the charge density
of poly(A) decreased with temperature, while that of poly(U) increased. These results are related to the distinct solution
structures of the RNA homopolymers. Considerable support was lent to this calorimetric approach by the excellent agreement
obtained in a test comparison between experimental and calculated parameters. From the intercept of energy term versus ln
[M+] plots, the non-electrostatic contributions, ΔH° and ΔS°, were determined. For each polynucleotide, the similarity in ΔG°
over the series of monovalent ions used in each study suggests a compensatory relationship between ΔH° and ΔS°, each of which
shows significant variation. The non-electrostatic contribution to binding of divalent magnesium is generally entropically
favorable and enthalpically unfavorable for both poly(A) and poly(U).
Received: 2 August 1995 / Accepted: 12 October 1995 相似文献
11.
Rate and equilibrium measurements of ryanodine binding to terminal cysternae fractions of heavy sarcoplasmic reticulum vesicles
demonstrate that its activation by high concentrations of monovalent salts is based on neither elevated osmolarity nor ionic
strength. The effect of the ions specifically depends on their chemical nature following the Hofmeister ion series for cations
(Li+ < NH+
4 < K−∼ Cs+≤ Na+) and anions (gluconate− < Cl− < NO3
−∼ ClO4
−∼ SCN−) respectively, indicating that both are involved in the formation of the salt-protein complex that can react with ryanodine.
Activation by rising salt concentrations exhibits saturation kinetics with different dissociation constants (25–11 m) and different degrees of cooperativity (n= 1.5–4.0) for the respective salts. Maximal second order binding rates between 40,000 and 80,000 (m
−1· sec−1) were obtained for chlorides and nitrates of 1a group alkali ions with the exception of lithium supporting only rates of
maximally 10,000 (M−1· sec−1). The nitrogen bases, NH+
4 and Tris+, in combination with chloride or nitrate, behave divergently. High maximal binding rates were achieved only with NH4NO3. The dissociation constants for the ryanodine–protein complexes obtained by measurements at equilibrium proved to depend
differently on salt concentration, yet, converging to 1–3 nm for the applied salts at saturating concentrations. The salts do not affect dissociation of the ryanodine protein complex
proving that the effect of salts on the protein's affinity for ryanodine is determined by their effect on the on-rate of ryanodine
binding. ATP and its analogues modify salt action resulting in elevated maximal binding rates and reduction or abolition of
binding cooperativity. Linear relations have been obtained by comparing the rates of ryanodine binding at different salt concentrations
with the rates or the initial amplitudes (15 sec) of salt induced calcium release from actively loaded heavy vesicles indicating
that the various salts promote specifically and concentration dependently channel opening and its reaction with ryanodine.
Received: 9 February 1998/Revised: 24 April 1998 相似文献
12.
Christoph Fertinger Alicja Franke Rudi van Eldik 《Journal of biological inorganic chemistry》2012,17(1):27-36
Compound I, an oxo–iron(IV) porphyrin π-cation radical species, and its one-electron-reduced form compound II are regarded as key intermediates in reactions catalyzed
by cytochrome P450. Although both reactive intermediates can be easily produced from model systems such as iron(III) meso-tetra(2,4,6-trimethylphenyl)porphyrin hydroxide by selecting appropriate reaction conditions, there are only a few thermal
activation parameters reported for the reactions of compound I analogues, whereas such parameters for the reactions of compound
II analogues have not been investigated so far. Our study demonstrates that ΔH
≠ and ΔS
≠ are closely related to the chemical nature of the substrate and the reactive intermediate (viz., compounds I and II) in epoxidation
and C–H abstraction reactions. Although most studied reactions appear to be enthalpy-controlled (i.e., ΔH
≠ > −TΔS
≠), different results were found for C–H abstractions catalyzed by compound I. Whereas the reaction with 9,10-dihydroanthracene
as a substrate is also dominated by the activation enthalpy (ΔH
≠ = 42 kJ/mol, ΔS
≠ = 41 J/Kmol), the same reaction with xanthene shows a large contribution from the activation entropy (ΔH
≠ = 24 kJ/mol, ΔS
≠ = −100 J/kmol). This is of special interest since the activation barrier for entropy-controlled reactions shows a significant
dependence on temperature, which can have an important impact on the relative reaction rates. As a consequence, a close correlation
between bond strength and reaction rate—as commonly assumed for C–H abstraction reactions—no longer exists. In this way, this
study can contribute to a proper evaluation of experimental and computational data, and to a deeper understanding of mechanistic
aspects that account for differences in the reactivity of compounds I and II. 相似文献
13.
O. Brix K. D. Clements R. M. G. Wells 《Journal of comparative physiology. B, Biochemical, systemic, and environmental physiology》1999,169(4-5):329-334
Haemoglobin components were analysed for nine species of New Zealand triplefins and their isoelectric points (pI) ranged
from 5.1 to 7.0. The number of well-expressed isohaemoglobins was larger in shallow-water and tidal pool species, ranging
from four in Grahamina signata to eight in Grahamina capito, and were relatively cathodal. Two strongly anodal isohaemoglobins were expressed in the mid-depth species Ruanoho decemdigitatus and Ruanoho whero, and one in the deeper water species Karalepis stewarti and Forsterygion malcolmi. The red blood cell oxygen-binding properties were determined at 15 °C and 25 °C in the pH range 6.7–7.9 for the shallow-water
species G. capito, the shallow to mid-depth species Forsterygion varium, and the deep-water species F. malcolmi. Oxygen affinity was highest for G. capito and the magnitude of the Bohr effect lower (Δlog P
50/ΔpH = −0.37 at 25 °C, where P
50 is the half-saturation coefficient) compared to the two Forsterygion species (Δlog P
50/ΔpH = −0.52 to −0.59). Further, the cooperativity factor, n
50, was lower in G. capito thus maintaining oxygen transport over a wide range of environmental oxygen pressures. Oxygen binding was similarly influenced
by temperature in both G. capito and F. malcolmi (maximum heat of oxygenation ΔHmax = −27 kJ mol−1 and −37 kJ mol−1, respectively). Thus, triplefin fishes living in shallow, thermally unstable habitats possess a greater number of cathodally
migrating isohaemoglobins, and their red blood cells have a higher oxygen affinity and reduced cooperativity which is less
sensitive to changes in pH than do species occurring in more stable, deeper water habitats. Our analysis of an assemblage
of closely related species circumvents some of the difficulties inherent in studies where interpretation of experimental results
is confounded by phylogeny.
Accepted: 18 March 1999 相似文献
14.
C. Immoos Michael G. Hill Donita Sanders James A. Fee Claire E. Slutter John H. Richards Harry B. Gray 《Journal of biological inorganic chemistry》1996,1(6):529-531
The electrochemistry of a water-soluble fragment from the CuA domain of Thermus thermophilus cytochrome ba
3
has been investigated. At 25 °C, CuA exhibits a reversible reduction at a pyridine-4-aldehydesemicarbazone-modified gold electrode (0.1 M Tris, pH 8) with E° = 0.24 V vs NHE. Thermodynamic parameters for the [Cu(Cys)2Cu]+/0 electrode reaction were determined by variable-temperature electrochemistry (ΔS°rc = –5.4(12) eu, ΔS° = –21.0(12) eu, ΔH° = –11.9(4) kcal/mol;ΔG° = –5.6 (11) kcal/mol). The relatively small reaction entropy is consistent with a low reorganization energy for [Cu(Cys)2Cu]+/0 electron transfer. An irreversible oxidation of [Cu(Cys)2Cu]+ at 1 V vs NHE confirms that the CuII:CuII state of CuA is significantly destabilized relative to the CuII state of analogous blue-copper proteins.
Received: 3 June 1996 / Accepted: 26 August 1996 相似文献
15.
Regeneration of transgenic plants of Mexican lime from Agrobacterium rhizogenes-transformed tissues 总被引:3,自引:0,他引:3
Transgenic Mexican lime [Citrus aurantifolia (Christm.) Swing] plants were regenerated from tissues transformed by Agrobacterium rhizogenes strain A4, containing the wild-type plasmid pRiA4 and the binary vector pESC4 with nos-npt II and cab-gus genes. Transgenic shoots were generated by two different approaches. The first approach used internodal stem segments cocultured
with A. rhizogenes. These were placed onto regeneration medium containing Murashige and Skoog salts and B5 organic compounds supplemented with
8 g ⋅ l–1 agar, 7.5 mg ⋅ l–1 6-benzylaminopurine, 1.0 mg ⋅ l–1 -naphthaleneacetic acid, 300 mg ⋅ l–1 cefotaxime and 80 mg ⋅ l–1 kanamycin as a selective agent, and incubated under continuous light at 25 °C. Under these conditions, 76% of the explants
produced shoots directly with no hairy root phase, with a mean of 1.3 shoots per explant, and 88% of these shoots were genetically
transformed as determined by β-glucuronidase (GUS) assays. In the second approach, segments of transformed roots (15 mm long) obtained from internodal stem
segments cocultured with A. rhizogenes were cultured on the above regeneration medium under similar conditions. Forty-one percent of these transformed root segments
produced adventitious shoots, with a mean of 2.2 shoots per explant and with 90% of shoots transformed. GUS activity was evident
in the transformed roots and in all parts of both transformed shoots and regenerated plants. The presence of the npt II and rolB genes in the regenerated plants was confirmed by PCR analysis. The presence of the npt II gene in the regenerated plants was also confirmed by Southern blot. Using these transformation systems, more than 300 Mexican
lime transgenic plants were obtained, 60 of which were adapted to growing in soil.
Received: 15 March 1997 / Revision received: 30 December 1997 / Accepted: 19 January 1998 相似文献
16.
S. C. J. Meskers C. Dennison Gerard W. Canters Harry P. J. M. Dekkers 《Journal of biological inorganic chemistry》1998,3(6):663-670
The dynamic quenching of the luminescence of racemic Eu(III)(pyridine-2,6-dicarboxylate=dpa)3
3– by the title proteins is investigated and the enantioselectivity of the proteins in the quenching of the Δ and Λ enantiomers
of Eu(dpa)3
3– is determined. The two diastereomeric quenching rate constants pertaining to azurin (k
q
Δ=3.3×106, k
q
Λ=2.7×106 M–1 s–1, pH 7.2, ionic strength I=22 mM) are lower than for its Met→44Lys mutant (k
q
Δ=1.9×107, k
q
Λ=1.4×107 M–1 s–1, same pH and I), indicating that energy transfer occurs from Eu(dpa)3
3– to the Cu(II) centre when the luminophore is bound to the hydrophobic patch of the protein near residue 44. The enantioselectivity
remains unaltered by the mutation: k
q
Δ/k
q
Λ=1.27±0.04, so Lys44 is probably not in direct contact with the Eu chelate. The I and pH dependence of k
q indicate that the lysine residue interacts electrostatically with Eu(dpa)3
3–. For plastocyanin the quenching rates are of the order of 106 M–1 s–1; for amicyanin they are two orders of magnitude larger (k
q
Δ=12×107, k
q
Λ=11×107 M–1 s–1, pH 7.2, I=22 mM). The variation of k
q is attributed to differences in the charge distribution on the proteins, which influences the binding of the luminophore
to the protein surface. For amicyanin the anion binding site near Lys59 and Lys60 may be involved in the energy transfer.
Received: 16 June 1998 / Accepted: 18 September 1998 相似文献
17.
Shaker B potassium channels undergo rapid N-type and slow C-type inactivation. While N-type inactivation is supposed to be mediated
by occlusion of the pore by the N-terminal protein structure, the molecular mechanisms leading to C-type inactivation are
less well understood. Considering N-type inactivation as a model for a protein conformational transition, we investigated
inactivation of heterologously expressed Shaker B potassium channels and mutants thereof, showing various degrees of C-type inactivation, under high hydrostatic (oil) pressure.
In addition to the derived apparent activation and reaction volumes (ΔV), experiments at various temperatures yielded estimates
for enthalpic (ΔH) and entropic (TΔS) contributions. N-type inactivation was accelerated by increasing temperature and slowed
by high hydrostatic pressure yielding at equilibrium ΔH = 76 kJ/mole, TΔS = 82 kJ/mole, and ΔV = 0.18 nm3 indicating that the transition to the N-type inactivated state is accompanied by an increase in volume and a decrease in
order. N-terminally deleted ShΔ6–46 constructs with additional mutations at either position 449 or 463 were used to investigate C-type inactivation. In
particular at high temperatures, inactivation occurred in two phases indicating more than one process. At equilibrium the
following values were estimated for the major inactivation component of mutant ShΔ6–46 T449A: ΔH = –64 kJ/mole, TΔS = –60 kJ/mole, and ΔV = –0.25 nm3, indicating that the C-type inactivated state occupies a smaller volume and is more ordered than the noninactivated state.
Thus, hydrostatic pressure affects N- and C-type inactivation in opposite ways.
Received: 17 May 1997 / Accepted: 18 July 1997 相似文献
18.
Timothy Darren Clark J. L. Rummer C. A. Sepulveda A. P. Farrell C. J. Brauner 《Journal of comparative physiology. B, Biochemical, systemic, and environmental physiology》2010,180(1):73-82
Tunas (family Scombridae) are exceptional among most teleost fishes in that they possess vascular heat exchangers which allow
heat retention in specific regions of the body (termed ‘regional heterothermy’). Seemingly exclusive to heterothermic fishes
is a markedly reduced temperature dependence of blood–oxygen (blood–O2) binding, or even a reversed temperature dependence where increasing temperature increases blood–O2 affinity. These unusual binding properties have been documented in whole blood and in haemoglobin (Hb) solutions, and they
are hypothesised to prevent oxygen loss from arteries to veins within the vascular heat exchangers and/or to prevent excessive
oxygen unloading to the warm tissues and ensure an adequate supply of oxygen to tissues positioned efferent to the heat exchangers.
The temperature sensitivity of blood–O2 binding has not been characterised in an ectothermic scombrid (mackerels and bonitos), but the existence of the unusual binding
properties in these fishes would have clear implications for their proposed association with regional heterothermy. Accordingly,
the present study examined oxygenation of whole blood of the chub mackerel (Scomber japonicus) at 10, 20 and 30°C and at 0.5, 1 and 2% CO2. Oxygen affinity was generally highest at 20°C for all levels of CO2. Temperature-independent binding was observed at low (0.5%) CO2, where the PO2 at 50% blood–O2 saturation (P
50) was not statistically different at 10 and 30°C (2.58 vs. 2.78 kPa, respectively) with an apparent heat of oxygenation (∆H°) close to zero (−6 kJ mol−1). The most significant temperature-mediated difference occurred at high (2%) CO2, where the P
50 at 10°C was twofold higher than that at 20°C with a corresponding ∆H° of +43 kJ mol−1. These results provide clear evidence of independent and reversed open-system temperature effects on blood oxygenation in
S. japonicus, and it is therefore speculated that these unusual blood–O2 binding characteristics may have preceded the evolution of vascular heat exchangers and regional heterothermy in fishes. 相似文献
19.
Marjon J. H. van Haandel I. M. C. M. Rietjens Ans E. M. F. Soffers Cees Veeger Jacques Vervoort Sandeep Modi Madhu S. Mondal Prasanta K. Patel Digambar V. Behere 《Journal of biological inorganic chemistry》1996,1(5):460-467
The second-order rate constants for the oxidation of a series of phenol derivatives by horseradish peroxidase compound II
were compared to computer-calculated chemical parameters characteristic for this reaction step. The phenol derivatives studied
were phenol, 4-chlorophenol, 3-hydroxyphenol, 3-methylphenol, 4-methylphenol, 4-hydroxybenzoate, 4-methoxyphenol and 4-hydroxybenzaldehyde.
Assuming a reaction of the phenolic substrates in their non-dissociated, uncharged forms, clear correlations (r = 0.977 and r = 0.905) were obtained between the natural logarithm of the second-order rate constants (ln k
app and ln k
2 respectively) for their oxidation by compound II and their calculated ionisation potential, i.e. minus the energy of their
highest occupied molecular orbital [E(HOMO)]. In addition to this first approach in which the quantitative structure-activity
relationship (QSAR) was based on a calculated frontier orbital parameter of the substrate, in a second and third approach
the relative heat of formation (ΔΔHF) calculated for the process of one-electron abstraction and H• abstraction from the phenol derivatives was used as a parameter. Plots of the natural logarithms of the second-order rate
constants (k
app and k
2) for the reaction and the calculated ΔΔHF values for the process of one-electron abstraction also provide clear QSARs with
correlation coefficients of –0.968 and –0.926 respectively. Plots of the natural logarithms of the second-order rate constants
(k
app and k
2) for the reaction and the calculated ΔΔHF values for the process of H• abstraction provide QSARs with correlation coefficients of –0.989 and –0.922 respectively. Since both mechanisms considered,
i.e. initial electron abstraction versus initial H• abstraction, provided clear QSARs, the results could not be used to discriminate between these two possible mechanisms for
phenol oxidation by horseradish peroxidase compound II. The computer calculation-based QSARs thus obtained for the oxidation
of the various phenol derivatives by compound II from horseradish peroxidase indicate the validity of the approaches investigated,
i.e. both the frontier orbital approach and the approach in which the process is described by calculated relative heats of
formation. The results also indicate that outcomes from computer calculations on relatively unrelated phenol derivatives can
be reliably compared to one another. Furthermore, as the actual oxidation of peroxidase substrates by compound II is known
to be the rate-limiting step in the overall catalysis by horseradish peroxidase, the QSARs of the present study may have implications
for the differences in the overall rate of substrate oxidation of the phenol derivatives by horseradish peroxidase.
Received: 29 March 1996 / Accepted: 17 July 1996 相似文献
20.
W Bode 《Journal of molecular biology》1979,127(4):357-374
p-Guanidinobenzoate-trypsinogen is transformed into a trypsin-like conformation upon binding of Ile-Val as evidenced by specific changes in its circular dichroism spectrum. By means of this signal the association constants for the binding of a variety of peptides sequentially analogous to either the bovine trypsin N-terminus or to the N-terminal activation peptide sequences of several trypsinogens have been determined at different Ca2+ concentrations. Ile-Val and Ile-Val-Gly exhibit the strongest binding affinity of all peptides investigated. Replacement of the first isoleucine or of the second valine residue by other amino acids considerably reduces the peptide affinity. Discussion of these is based on the known spatial arrangement of the Ile16-Val17-Gly18 N-terminus and of the Ile-Val dipeptide in the Ile16 cleft (crystal structures of bovine trypsin and of the trypsinogen-PTI3-Ile-Val complex; Bode et al., 1978). The free energies of binding of the first and of the second peptide residue are almost additive indicating independency between both subsites. The third residue, glycine, does not significantly contribute to binding. The peptide analogues of various trypsinogen N-termini exhibit no measurable affinity for the Ile 16 cleft.The equilibrium constant for the binding of PTI to trypsinogen and the affinity of Ile-Val for the resulting binary complex have been determined in the presence and absence of Ca2+, using the competitive PTI-binding to α-chymotrypsin. These competition experiments allow the estimation of the standard free-energy changes due to the conformational transition of trypsinogen into a trypsin-like state (+43 kJ mol?1, 20 °C; stabilization of the “activation domain”; Fehlhammer et al., 1977), due to the binding of the trypsin N-terminus (—55 kJ mol?1) and of the peptide analogues (e.g. Ile-Val; ?28 kJ mol?1) into the preformed Ile 16 cleft, and due to the specific burying of the covalently linked pGB group in the fixed specificity pocket (— 39 kJ mol?1). This pocket is co-operatively linked with the Ile 16 cleft according to a free-energy change coupling of —43 kJ mol?1. 相似文献