Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Progress-curve equations for reversible enzyme-catalysed reactions inhibited by tight-binding inhibitors.

ABSTRACT: The rate equation for a tight-binding inhibitor of an enzyme-catalysed first-order reversible reaction was used to derive two integrated equations. One of them covers the situations in which competitive, uncompetitive or non-competitive inhibition occurs and the other refers to the special non-competitive case where the two inhibition constants are equal. For these equations, graphical and non-linear regression methods are proposed for distinguishing between types of inhibition and for calculating inhibition constants from progress-curve data. The application of the non-linear regression to the analysis of stimulated progress curves in the presence of a tight-binding inhibitor is also presented. The results obtained are valid for any type of 'dead-end'-complex-forming inhibitor and can be used to characterize an unknown inhibitor on the basis of progress curves.

SUBMITTER: Szedlacsek SE

PROVIDER: S-EPMC1133683 | biostudies-other | 1990 Feb

REPOSITORIES: biostudies-other

ACCESS DATA

Json Xml

Similar Datasets

Progress-curve analysis in enzyme kinetics. Numerical solution of integrated rate equations.

Project description:Progress curves of enzyme-catalysed reactions are described by equations of a type that precludes direct calculation of the extent of reaction at any time. Previously, such equations have been solved by the Newton-Raphson method, but this procedure may fail when based upon the usual formulae. An alternative formulation is proposed that is both quicker and more robust.

| S-EPMC1146728 | biostudies-other

Analysis of progress curves for enzyme-catalysed reactions. Automatic construction of computer programs for fitting integrated rate equations.

Project description:The computer analysis of progress curves for enzyme-catalysed reactions involves a series of mathematical and computational tasks. The three most daunting of these are the derivation of an integrated rate equation, solving this equation so that the amount of product formed by the reaction at any time can be calculated, and incorporating this solution into a non-linear-regression computer program. This paper describes the basis of a computer program that greatly simplifies the problem. The proposed mechanism is specified in the familiar kinetic constant form, which is automatically translated into a program capable of fitting this mechanism to a series of experimental progress curves. The approach is illustrated for a reversible reaction with one substrate and one product, and tested with some data obtained for the fumarase reaction. A copy of the program has been deposited as Supplementary Publication SUP 50148 (13 pages) at the British Library Document Supply Centre, Boston Spa, Wetherby, West Yorkshire LS23 7BQ, U.K., from whom copies can be obtained on the terms indicated in Biochem. J. (1989) 257, 5.

| S-EPMC1138375 | biostudies-other

Integrated rate equations for enzyme-catalysed first-order and second-order reactions.

Project description:Generalized rate equations covering all mechanisms giving hyperbolic initial-rate kinetics with stoichiometry A in equilibrium P, A in equilibrium P + Q, A + B in equilibrium P and A + B in equilibrium P + Q were integrated. The results are regular and reasonably economical.

| S-EPMC1144258 | biostudies-other

Integrated rate equations for irreversible enzyme-catalysed first-order and second-order reactions.

Project description:Integrated rate equations are presented that describe irreversible enzyme-catalysed first-order and second-order reactions. The equations are independent of the detailed mechanism of the reaction, requiring only that it be hyperbolic and unbranched. The results should be directly applicable in the laboratory.

| S-EPMC1144673 | biostudies-other

Dynamical origins of heat capacity changes in enzyme-catalysed reactions.

Project description:Heat capacity changes are emerging as essential for explaining the temperature dependence of enzyme-catalysed reaction rates. This has important implications for enzyme kinetics, thermoadaptation and evolution, but the physical basis of these heat capacity changes is unknown. Here we show by a combination of experiment and simulation, for two quite distinct enzymes (dimeric ketosteroid isomerase and monomeric alpha-glucosidase), that the activation heat capacity change for the catalysed reaction can be predicted through atomistic molecular dynamics simulations. The simulations reveal subtle and surprising underlying dynamical changes: tightening of loops around the active site is observed, along with changes in energetic fluctuations across the whole enzyme including important contributions from oligomeric neighbours and domains distal to the active site. This has general implications for understanding enzyme catalysis and demonstrating a direct connection between functionally important microscopic dynamics and macroscopically measurable quantities.

| S-EPMC5862990 | biostudies-literature

SAM-dependent enzyme-catalysed pericyclic reactions in natural product biosynthesis.

Project description:Pericyclic reactions-which proceed in a concerted fashion through a cyclic transition state-are among the most powerful synthetic transformations used to make multiple regioselective and stereoselective carbon-carbon bonds. They have been widely applied to the synthesis of biologically active complex natural products containing contiguous stereogenic carbon centres. Despite the prominence of pericyclic reactions in total synthesis, only three naturally existing enzymatic examples (the intramolecular Diels-Alder reaction, and the Cope and the Claisen rearrangements) have been characterized. Here we report a versatile S-adenosyl-l-methionine (SAM)-dependent enzyme, LepI, that can catalyse stereoselective dehydration followed by three pericyclic transformations: intramolecular Diels-Alder and hetero-Diels-Alder reactions via a single ambimodal transition state, and a retro-Claisen rearrangement. Together, these transformations lead to the formation of the dihydropyran core of the fungal natural product, leporin. Combined in vitro enzymatic characterization and computational studies provide insight into how LepI regulates these bifurcating biosynthetic reaction pathways by using SAM as the cofactor. These pathways converge to the desired biosynthetic end product via the (SAM-dependent) retro-Claisen rearrangement catalysed by LepI. We expect that more pericyclic biosynthetic enzymatic transformations remain to be discovered in naturally occurring enzyme 'toolboxes'. The new role of the versatile cofactor SAM is likely to be found in other examples of enzyme catalysis.

| S-EPMC5679075 | biostudies-literature

Estimation of the initial velocity of enzyme-catalysed reactions by non-linear regression analysis of progress curves.

Project description:Most methods for studying the kinetic properties of an enzyme involve the determination of initial velocities. When the reaction progress curve shows significant curvature due to depletion of the substrate, accumulation of inhibitory products or instability of the enzyme, estimation of the initial velocity is a subjective and inexact process. Two methods have been suggested [Cornish-Bowden (1975) Biochem. J. 144, 305-312; Boeker (1982) Biochem J. 203, 117-123] that attempt to eliminate this subjective element. The present study offers a third alternative, which is based on fitting a reparameterized form of the integrated Michaelis-Menten equation to the progress curves by non-linear regression. This method yields estimates and standard errors of the initial velocity and of the time to reach 50% reaction. No prior knowledge of the apparent product concentration at zero time or infinite time is required, since both of these quantities are also estimated from the data. It is shown that this method yields reliable estimates of the initial velocity under a wide range of circumstances, including those where the two previously published methods perform poorly.

| S-EPMC1144952 | biostudies-other

Hydrogen tunnelling in enzyme-catalysed H-transfer reactions: flavoprotein and quinoprotein systems.

Project description:It is now widely accepted that enzyme-catalysed C-H bond breakage occurs by quantum mechanical tunnelling. This paradigm shift in the conceptual framework for these reactions away from semi-classical transition state theory (TST, i.e. including zero-point energy, but with no tunnelling correction) has been driven over the recent years by experimental studies of the temperature dependence of kinetic isotope effects (KIEs) for these reactions in a range of enzymes, including the tryptophan tryptophylquinone-dependent enzymes such as methylamine dehydrogenase and aromatic amine dehydrogenase, and the flavoenzymes such as morphinone reductase and pentaerythritol tetranitrate reductase, which produced observations that are also inconsistent with the simple Bell-correction model of tunnelling. However, these data-especially, the strong temperature dependence of reaction rates and the variable temperature dependence of KIEs-are consistent with other tunnelling models (termed full tunnelling models), in which protein and/or substrate fluctuations generate a configuration compatible with tunnelling. These models accommodate substrate/protein (environment) fluctuations required to attain a configuration with degenerate nuclear quantum states and, when necessary, motion required to increase the probability of tunnelling in these states. Furthermore, tunnelling mechanisms in enzymes are supported by atomistic computational studies performed within the framework of modern TST, which incorporates quantum nuclear effects.

| S-EPMC1647315 | biostudies-literature

Measurement of Net Rate Constants from Enzyme Progress Curves without Curve Fitting.

Project description:A method is described whereby net rate constants can be directly inferred from the progress curves of enzyme intermediates without the need for model specification, numerical analysis, curve fitting, or the steady-state approximation. Specifically, if an enzyme intermediate in an ultimately irreversible serial subsequence is perturbed from and returns back to its equilibrium state as the substrate is consumed, then its net rate constant is given by the ratio of the total substrate consumed and the area under the progress curve for the enzyme intermediate. A rigorous analysis demonstrates this result to hold independent of the complete enzymatic reaction in which the subsequence is embedded, making it broadly applicable to a very wide range of kinetic mechanisms, including those complicated by inhibition. As a theoretical consequence, it is shown that traditionally steady-state parameters such as kcat, kcat/KM, and net rate constants can be expressed as limiting ratios of averages without requiring the steady-state hypothesis. Finally, a mock data set is generated for a system of contemporary interest that can serve as both an example of how the methodology would be used in practice and a proof of concept.

| S-EPMC6910778 | biostudies-literature

Real-Time Monitoring of Enzyme-Catalysed Reactions using Deep UV Resonance Raman Spectroscopy.

Project description:For enzyme-catalysed biotransformations, continuous in situ detection methods minimise the need for sample manipulation, ultimately leading to more accurate real-time kinetic determinations of substrate(s) and product(s). We have established for the first time an on-line, real-time quantitative approach to monitor simultaneously multiple biotransformations based on UV resonance Raman (UVRR) spectroscopy. To exemplify the generality and versatility of this approach, multiple substrates and enzyme systems were used involving nitrile hydratase (NHase) and xanthine oxidase (XO), both of which are of industrial and biological significance, and incorporate multistep enzymatic conversions. Multivariate data analysis of the UVRR spectra, involving multivariate curve resolution-alternating least squares (MCR-ALS), was employed to effect absolute quantification of substrate(s) and product(s); repeated benchmarking of UVRR combined with MCR-ALS by HPLC confirmed excellent reproducibility.

| S-EPMC5488198 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data