scholarly journals Photochemical and ligand-exchange properties of the cyanide complex of fully reduced cytochrome c oxidase

1991 ◽  
Vol 279 (2) ◽  
pp. 355-360 ◽  
Author(s):  
B C Hill ◽  
S Marmor
Keyword(s):  
Binding Site ◽  
Cytochrome Oxidase ◽  
Ligand Exchange ◽  
Binding Constant ◽  
First Order ◽  
Structure And Dynamics ◽  
Equilibrium Binding ◽  
Recombination Kinetics ◽  
Equilibrium Binding Constant ◽  
Reduced State

Cytochrome oxidase, in its fully reduced state, forms a complex with CN having a Kd of 230 microM with a stoicheiometry of 1 CN molecule per cytochrome oxidase. We do not detect a second CN-binding site as seen by i.r. spectroscopy [Yoshikawa & Caughey (1990) J. Biol. Chem. 265, 7945-7958]. The ferrocytochrome a3-CN complex, like the analogous ferrocytochrome a3-CO complex, is photosensitive but with a 15-fold lower quantum yield for photolysis. Analysis of the recombination kinetics after CN photolysis establishes a simple bimolecular binding constant of 235 M-1.s-1, in agreement with the value obtained from stopped-flow studies [Antonini, Brunori, Greenwood, Malmström & Rotillo (1971) Eur. J. Biochem. 23, 396-400]. A rate of 0.07 s-1 for the first-order dissociation of CN from cytochrome a3 is found by the rate of exchange of CO with ferrocytochrome a3-CN, and is consistent with the value calculated from the equilibrium binding constant and the CN on rate. However, O2 is able to oxidize the fully reduced CN compound at a rate well in excess of the CN off rate. The product of this oxidation reaction is a partially reduced CN complex. This implies that O2 either promotes CN dissociation or is able to oxidize the CN-bound enzyme directly. These results are discussed in the context of the structure and dynamics of the ligand-binding site of cytochrome oxidase.

Does the ligand-biopolymer equilibrium binding constant depend on the number of bound ligands?

Biopolymers ◽  
2010 ◽  
Vol 93 (11) ◽  
pp. 932-935 ◽  
Author(s):  
Daria A. Beshnova ◽  
Anastasia O. Lantushenko ◽  
Maxim P. Evstigneev
Keyword(s):  
Binding Constant ◽  
Equilibrium Binding ◽  
Equilibrium Binding Constant

scholarly journals The linkage between binding of the C-terminal domain of hirudin and amidase activity in human α-thrombin

1993 ◽  
Vol 289 (2) ◽  
pp. 475-480 ◽  
Author(s):  
R de Cristofaro ◽  
B Rocca ◽  
B Bizzi ◽  
R Landolfi
Keyword(s):  
Rate Constants ◽  
Binding Constant ◽  
Amidase Activity ◽  
Equilibrium Binding ◽  
Viscosity Effects ◽  
The Individual ◽  
Individual Rate ◽  
Hydrolysis Of ◽  
Equilibrium Binding Constant

A method derived from the analysis of viscosity effects on the hydrolysis of the amide substrates D-phenylalanylpipecolyl-arginine-p-nitroaniline, tosylglycylprolylarginine-p-nitroanaline and cyclohexylglycylalanylarginine-p-nitroalanine by human alpha-thrombin was developed to dissect the Michaelis-Menten parameters Km and kcat into the individual rate constants of the binding, acylation and deacylation reactions. This method was used to analyse the effect of the C-terminal hirudin (residues 54-65) [hir-(54-65)] domain on the binding and hydrolysis of the three substrates. The results showed that the C-terminal hir-(54-65) fragment affects only the acylation rate, which is increased approx. 1.2-fold for all the substrates. Analysis of the dependence of acylation rate constants on hirudin-fragment concentration, allowed the determination of the equilibrium binding constant of C-terminal hir-(54-65) (Kd approximately 0.7 microM). In addition this peptide was found to competitively inhibit thrombin-fibrinogen interaction with a Ki which is in excellent agreement with the equilibrium constant derived from viscosity experiments. These results demonstrate that binding of hir-(54-65) to the fibrinogen recognition site of thrombin does not affect the equilibrium binding of amide substrates, but induces only a small increase in the acylation rate of the hydrolysis reaction.

scholarly journals A comparison of acylation, phosphorylation and binding in related substrates and inhibitors of serum cholinesterase

1966 ◽  
Vol 101 (3) ◽  
pp. 584-590 ◽  
Author(s):  
A R Main ◽  
F L Hastings
Keyword(s):  
Rate Constant ◽  
Binding Constant ◽  
Binding Energies ◽  
Serum Cholinesterase ◽  
P Values ◽  
Ethyl Methyl ◽  
Equilibrium Binding ◽  
Alkyl Groups ◽  
Inhibition Constants ◽  
Equilibrium Binding Constant

1. The K(m) and catalytic-centre activities for human serum cholinesterase and methyl, ethyl, n-propyl and n-butyl butyrate substrates were determined and compared with the related inhibition constants of a similarly substituted organophosphate inhibitor series based on malaoxon. The results indicated that the catalytic-centre activities approximated to k(+2(a)), the acylation rate constant, and that K(m) approximated to the equilibrium binding constant. The inhibition constants measured were K(a), the equilibrium binding constant, and k(+2(p)), the phosphorylation rate constant. 2. The effects of the alkyl substituents on k(+2(p)) and k(+2(a)) were closely parallel, and the decreasing order in each case was: n-butyl; methyl; n-propyl; ethyl. The Taft constants did not follow this order, suggesting that alkyl substituents did not primarily effect acylation or phosphorylation by electron induction. 3. For comparable homologues, the k(+2(a)) values were on average 435 times the k(+2(p)) values. The k(+2(p)) values at 25 degrees and pH7.6 ranged from 6.6min.(-1) for the diethyl member to 22.6min.(-1) for the di-n-butyl member. 4. The effect of the alkyl substituents on K(a) and K(m) were closely paralleled. The increasing order in each case was: n-butyl; n-propyl; ethyl; methyl. The K(a) values were about 100 times less than the comparable K(m) values. 5. Consideration of the binding energies suggested that only one of the two alkyl groups on the malaoxon homologues bound to the active site. 6. The possibility that malaoxon acted as a substrate as well as an inhibitor for cholinesterase was also investigated, but no evidence of a substrate reaction was found.

scholarly journals Architecture of Allosteric Structure. Rate Equations, Rate Constants, and Equilibrium Constants for Reaction of: Hb4 with O2 and (HbO2)4 with Dithionate, in the Presence of 2,3-Bisphosphoglycerate

2019 ◽  
Author(s):  
Francis Knowles ◽  
Samantha J. Doyle ◽  
Douglas Magde
Keyword(s):  
Rate Constants ◽  
Binding Constant ◽  
Equilibrium Constants ◽  
Rate Equations ◽  
Rate Law ◽  
First Order ◽  
Equilibrium Binding ◽  
Binding Curve ◽  
O2 Binding

Three unknown quantities are all that is required to describe the O2-equilibrium binding curve for fractional saturation of human hemoglobin in red blood cells, under standard conditions: Kα, the O2-binding constant of equivalent α-chains; KC, the equilibrium constant for the T →R conformation change; Kβ, the O2-binding constant of equivalent β-chains. The model for formulation of the equation of state is a 3-stage ordered sequence of reactions. The values of were established by determination of rate constants for the oxygenation reaction and the dithionite-mediated de oxygenation reaction. The rate law for the forward reaction in the presence of excess O2 yields The same rate law yields for the dithionite-mediated de-oxygenation reaction. The rate constants for binding O2 are pseudo-first-order. The rate constants for release of O2 are first-order. Reactions involving O2, are 2-step ordered sequences of equivalent subunits. Progress curves for a 2-step ordered sequence of equivalent chains collapse to a first order reaction. Progress curves for both oxygenation and dithionite-mediated de-oxygenation reactions return is 0.0580 for the oxygenation reaction and 0.0358 for the dithionite-mediated de-oxygenation reaction. The corresponding values from the O2-equilibrium binding curve are: and = 0.02602. Values of determined from rate constants of progress curves for oxygenation and dithionite-mediated de-oxygenation reactions are close to values of determined by analysis of the O2-equilibrium binding curves for whole blood, by the Perutz/Adair equation.<br>

scholarly journals Architecture of Allosteric Structure. Rate Equations, Rate Constants, and Equilibrium Constants for Reaction of: Hb4 with O2 and (HbO2)4 with Dithionate, in the Presence of 2,3-Bisphosphoglycerate

2019 ◽  
Author(s):  
Francis Knowles ◽  
Samantha J. Doyle ◽  
Douglas Magde
Keyword(s):  
Rate Constants ◽  
Binding Constant ◽  
Equilibrium Constants ◽  
Rate Equations ◽  
Rate Law ◽  
First Order ◽  
Equilibrium Binding ◽  
Binding Curve ◽  
O2 Binding

Three unknown quantities are all that is required to describe the O2-equilibrium binding curve for fractional saturation of human hemoglobin in red blood cells, under standard conditions: Kα, the O2-binding constant of equivalent α-chains; KC, the equilibrium constant for the T →R conformation change; Kβ, the O2-binding constant of equivalent β-chains. The model for formulation of the equation of state is a 3-stage ordered sequence of reactions. The values of were established by determination of rate constants for the oxygenation reaction and the dithionite-mediated de oxygenation reaction. The rate law for the forward reaction in the presence of excess O2 yields The same rate law yields for the dithionite-mediated de-oxygenation reaction. The rate constants for binding O2 are pseudo-first-order. The rate constants for release of O2 are first-order. Reactions involving O2, are 2-step ordered sequences of equivalent subunits. Progress curves for a 2-step ordered sequence of equivalent chains collapse to a first order reaction. Progress curves for both oxygenation and dithionite-mediated de-oxygenation reactions return is 0.0580 for the oxygenation reaction and 0.0358 for the dithionite-mediated de-oxygenation reaction. The corresponding values from the O2-equilibrium binding curve are: and = 0.02602. Values of determined from rate constants of progress curves for oxygenation and dithionite-mediated de-oxygenation reactions are close to values of determined by analysis of the O2-equilibrium binding curves for whole blood, by the Perutz/Adair equation.<br>

scholarly journals Applying support-vector machine learning algorithms toward predicting host–guest interactions with cucurbit[7]uril

2020 ◽  
Vol 22 (26) ◽  
pp. 14976-14982
Author(s):  
Anthony Tabet ◽  
Thomas Gebhart ◽  
Guanglu Wu ◽  
Charlie Readman ◽  
Merrick Pierson Smela ◽  
...  
Keyword(s):  
Machine Learning ◽  
Support Vector Machine ◽  
Support Vector Machines ◽  
Small Molecules ◽  
Binding Constant ◽  
Machine Learning Algorithms ◽  
Support Vector ◽  
Equilibrium Binding ◽  
Vector Machines ◽  
Equilibrium Binding Constant

We evaluate the ability of support-vector machines to predict the equilibrium binding constant of small molecules to cucurbit[7]uril.

Antithrombin-mediated Inhibition of Factor Vlla-Tissue Factor Complex by the Synthetic Pentasaccharide Representing the Heparin Binding Site to Antithrombin

1996 ◽  
Vol 76 (01) ◽  
pp. 005-008 ◽  
Author(s):  
Jean Claude Lormeau ◽  
Jean Pascal Herault ◽  
Jean Marc Herbert
Keyword(s):  
Binding Site ◽  
Tissue Factor ◽  
Residual Activity ◽  
Coagulation Factors ◽  
Heparin Binding ◽  
Experimental Conditions ◽  
First Order ◽  
Heparin Binding Site ◽  
Coagulant Activity

SummaryWe examined the effect of the synthetic pentasaccharide representing the minimal binding site of heparin to antithrombin on the antithrombin-mediated inactivation of factor Vila bound to tissue factor. This effect was compared to the effect of unfractionated heparin. Using purified recombinant human coagulation factors and either a clotting or an amidolytic assay for the determination of the residual activity of factor Vila, we showed that the pentasaccharide was an efficient antithrombin-dependent inhibitor of the coagulant activity of tissue factor-factor Vila complex. In our experimental conditions, assuming a mean MW of 14,000 for heparin, the molar pseudo-first order rate constants for ATIII-mediated FVIIa inhibition by ATIII-binding heparin and by the synthetic pentasaccharide were found to be similar with respective values of 104,000 ± 10,500 min-1 and 112,000 ± 12,000 min-1 (mean ± s.e.m., n = 3)

Enhancement of Plasminogen Binding to U937 Cells and Fibrin by Complestatin

1997 ◽  
Vol 77 (01) ◽  
pp. 137-142 ◽  
Author(s):  
Kiyoshi Tachikawa ◽  
Keiji Hasurni ◽  
Akira Endo
Keyword(s):  
Binding Site ◽  
Binding Affinity ◽  
Blood Cells ◽  
Aminocaproic Acid ◽  
U937 Cells ◽  
Biochemical Basis ◽  
Equilibrium Binding ◽  
Pharmacological Stimulation ◽  
Endogenous Fibrinolysis ◽  
Stimulation Of

SummaryPlasminogen binds to endothelial and blood cells as well as to fibrin, where the zymogen is efficiently activated and protected from inhibition by α2-antiplasmin. In the present study we have found that complestatin, a peptide-like metabolite of a streptomyces, enhances binding of plasminogen to cells and fibrin. Complestatin, at concentrations ranging from 1 to 5 μM, doubled 125I-plasminogen binding to U937 cells both in the absence and presence of lipoprotein(a), a putative physiological competitor of plasminogen. The binding of 125I-plasminogen in the presence of complestatin was abolished by e-aminocaproic acid, suggesting that the lysine binding site(s) of the plasminogen molecule are involved in the binding. Equilibrium binding analyses indicated that complestatin increased the maximum binding of 125I-plasminogen to U937 cells without affecting the binding affinity. Complestatin was also effective in increasing 125I-plasminogen binding to fibrin, causing 2-fold elevation of the binding at ~1 μM. Along with the potentiation of plasminogen binding, complestatin enhanced plasmin formation, and thereby increased fibrinolysis. These results would provide a biochemical basis for a pharmacological stimulation of endogenous fibrinolysis through a promotion of plasminogen binding to cells and fibrin.

The Orientation of Cytochrome c Oxidase in Coupled Phospholipid Membrane Vesicles—A Spin-Label Study

1975 ◽  
Vol 53 (3) ◽  
pp. 364-370 ◽  
Author(s):  
J. A. Kornblatt ◽  
W. L. Chen ◽  
J. C. Hsia ◽  
G. R. Williams
Keyword(s):  
Molecular Weight ◽  
Binding Site ◽  
Cytochrome Oxidase ◽  
Spin Label ◽  
Resonance Spectrum ◽  
Membrane Vesicles ◽  
Phospholipid Membrane ◽  
Vesicle Membrane ◽  
Exterior Surface ◽  
Label Study

Cytochrome oxidase, an enzyme containing six different subunits, has been shown to span the inner mitochrondrial membrane. The arrangement of the subunits within the membrane is unknown. We have specifically labeled the 25 000 molecular weight subunit with a spin-label derivative of N-ethylmaleimide, 3-maleimido-2,2,5,5-tetramethyl-1-pyrrolidinyloxyl (NEM-SL(5)). NEM-SL(5)-labeled cytochrome oxidase can be incorporated into phospholipid membranes to form coupled vesicles of the Hinkle, Kim &Racker ((1972) J. Biol. Chem. 247, 1338–1339) type. The resonance spectrum of NEM-SL(5) is similar in both soluble and vesicular cytochrome oxidase. Since ascorbate has been shown to reduce only spin label that is exposed to the exterior surface of a closed vesicle, we have used ascorbate to determine the NEM-SL(5)-binding site in the coupled vesicles. NEM-SL(5)-labeled cytochrome oxidase vesicles are reduced by 10 mM ascorbate with [Formula: see text] of 1 min at 22 °C. The rate of reduction is relatively independent of temperature. We conclude that (1) cytochrome oxidase is unidirectionally or preferentially oriented in the vesicle membrane, and (2) the NEM-SL(5)-binding site on the 25 000 molecular weight subunit is exposed to the external aqueous medium.

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