Name: graphing enzyme kinetics data
Uploaded: 2023-04-30T10:09:42.000Z
Duration: 1 min 19 s
Channel: JoVE

K12 Standards

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North Carolina K-12 Science Standard Course of Study

9-12

Biology

from-molecules-to-organisms-structures-and-processes

From Molecules to Organisms- Structures and Processes

LS.Bio.1.2

Introduction to Enzyme Kinetics

Enzyme kinetics studies the rates of enzyme-catalyzed reactions. The rates of repeated experiments at varying substrate concentrations are monitored by measuring the concentration of substrate consumed or product formed over time.
These results can be graphed to show how substrate concentration affects the rate or velocity of a reaction.
The reaction velocity increases linearly with increasing substrate amounts at low concentrations, but it begins to plateau at higher concentrations. The rate approaches a maximum velocity or Vmax &#8211; the rate where the enzyme is completely saturated with the substrate.
Enzyme affinity measures how strongly or weakly an enzyme binds its substrate and is quantified by KM, the Michaelis constant. The value of KM is equal to the substrate concentration when the rate is 50% of the Vmax.
A small KM indicates an enzyme has a high substrate affinity and vice versa. An enzyme with a larger KM requires higher substrate concentrations to approach its maximum velocity compared to an enzyme with a lower KM.

inleiding tot enzymkinetiek

Enzymkinetiek bestudeert de snelheid van biochemische reacties. Wetenschappers monitoren de reactiesnelheden voor een bepaalde enzymatische reactie bij verschillende substraatconcentraties. Er kunnen ook aanvullende onderzoeken worden uitgevoerd met remmers of andere moleculen die de reactiesnelheid beïnvloeden.

De onderzoeker kan vervolgens de initiële reactiesnelheid of snelheid (Vo) van een bepaalde proef uitzetten tegen de substraatconcentratie ([S]) om een grafiek van de reactie-eigenschappen te verkrijgen. Voor veel enzymatische reacties waarbij één enkel substraat betrokken is, passen deze gegevens in de Michaelis-Menten-vergelijking, een vergelijking afgeleid door Leonor Michaelis en Maud Menten.

<img alt="Eq1" src="/files/ftp_upload/12164/12164_Equation 1.png" style="text-align: center; height: 70px;" />

De vergelijking schat de maximale snelheid (Vmax) en de Michaelis-constante (KM) voor het onderzochte enzym en is gebaseerd op de volgende aannames:

<ol>
	<li>Bij het begin van de reactie is er geen product aanwezig.</li>
	<li>De snelheid van de vorming van enzym-substraatcomplexen is gelijk aan de snelheid van dissociatie en afbraak in producten.</li>
	<li>De enzymconcentratie is minimaal vergeleken met de substraatconcentratie.</li>
	<li>Alleen de initiële reactiesnelheden worden gemeten.</li>
	<li>Het enzym is aanwezig in de vrije vorm of in het enzym-substraatcomplex.</li>
</ol>

Verschillende herschikkingen van de Michaelis-Menten-vergelijking, zoals de Lineweaver-Burke-, Eadie-Hofsteot- en Hanes-Woolf-plots, zijn alternatieve manieren om kinetische parameters in een grafiek weer te geven. De Lineweaver-Burke of dubbele reciproke plot wordt vaak gebruikt om de KM en de Vmax te schatten. De grafiek gebruikt de omgekeerde waarden van de x- en y-as uit de Michaelis-Menten-grafiek. Wiskundig gezien is het y-snijpunt gelijk aan 1/Vmax, en het x-snijpunt gelijk aan −1/KM.

De Lineweaver-Burke-plot kan worden gebruikt om visueel onderscheid te maken tussen typen remmers: competitief, niet-competitief en niet-competitief. Verschillende herschikkingen van de Michaelis-Menten-vergelijking, zoals de Eadie-Hofstee- en Hanes-Woolf-plots, worden ook gebruikt om kinetische parameters te bepalen.

Inleiding tot enzymkinetiek

Enzyme kinetics studies the rates of biochemical reactions. Scientists monitor the reaction rates for a particular enzymatic reaction at various substrate ...

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JoVE Core

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KEY TERMS AND CONCEPTS

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graphing enzyme kinetics data

Enzyme kinetics uses graphs to show how fast a biochemical reaction happens. Scientists measure the initial reaction rate, or velocity (Vo), at different substrate concentrations ([S]). They may also run trials with inhibitors or other molecules that change the reaction rate.

The results are often plotted as Vo against [S]. For many single-substrate reactions, the data fits the Michaelis-Menten equation. This equation, named after Leonor Michaelis and Maud Menten, helps estimate the maximum velocity (Vmax) and the Michaelis constant (KM) for the enzyme being studied.

The Michaelis-Menten model is based on a few assumptions. No product is present at the start of the reaction. The rate of enzyme-substrate complex formation equals the rate of its breakdown into products and its dissociation. The enzyme concentration is minimal compared with the substrate concentration. The model also uses only the initial reaction rates. In this setup, the enzyme is either free or bound in an enzyme-substrate complex.

Scientists can also use other graph forms to analyze the same data. The Lineweaver-Burke plot, also called the double reciprocal plot, uses the reciprocal values from the Michaelis-Menten graph. On this plot, the y-intercept equals 1/Vmax and the x-intercept equals -1/KM. It is often used to estimate KM and Vmax and to visually distinguish competitive, non-competitive, and uncompetitive inhibitors.

Other rearrangements of the Michaelis-Menten equation include the Eadie-Hofstee plot and the Hanes-Woolf plot. These graphs are also used to determine kinetic parameters. Together, these methods help scientists compare reaction behavior and understand how enzymes respond to substrate concentration and inhibitors.

Graphing Enzyme Kinetics Data

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Learn how enzyme kinetics data is graphed with Michaelis-Menten, Lineweaver-Burke, Eadie-Hofstee, and Hanes-Woolf plots to estimate Vmax and KM.

Graphing Enzyme Kinetics Data | JoVE K12

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HIgh School

BIO.2

BIO.2.c

Chemistry

CH.7

CH.7.f

Florida Department of Education

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Graphing Enzyme Kinetics Data

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