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

K12 Standards

north-carolina-k-12-science-standard-course-of-study

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

La cinética enzimática estudia las tasas de reacciones catalizadas por enzimas. Las tasas de experimentos repetidos a diferentes concentraciones de sustrato se monitorean midiendo la concentración de sustrato consumido o producto formado a lo largo del tiempo.
Estos resultados se pueden graficar para mostrar cómo la concentración de sustrato afecta la velocidad o la velocidad de una reacción.
La velocidad de reacción aumenta linealmente con el aumento de las cantidades de sustrato a bajas concentraciones, pero comienza a estabilizarse a concentraciones más altas. La velocidad se acerca a una velocidad máxima o Vmáx., la velocidad en la que la enzima está completamente saturada con el sustrato.
La afinidad enzimática mide la fuerza o debilidad con la que una enzima se une a su sustrato y se cuantifica mediante KM, la constante de Michaelis. El valor de KM es igual a la concentración de sustrato cuando la tasa es el 50% de la Vmáx.
Un KM pequeño indica que una enzima tiene una alta afinidad por el sustrato y viceversa. Una enzima con un KM mayor requiere concentraciones de sustrato más altas para acercarse a su velocidad máxima en comparación con una enzima con un KM más bajo.

<ol>
	<li>Voet D. y Voet J.G. Biochemistry, 4ª ed. John Wiley & Sons, 2010 pp-487-492</li>
	<li>Berg J.M., Tymoczko J.L. y Stryer L. Bioquímica, 5ª ed., W.H. Freeman and Co., Sección 8.4</li>
</ol>

introducción a la cinética enzimática

La cinética enzimática estudia las velocidades de las reacciones bioquímicas. Los científicos monitorean las velocidades de reacción de una reacción enzimática particular en varias concentraciones de sustrato. También se pueden realizar pruebas adicionales con inhibidores u otras moléculas que afecten la velocidad de reacción.

Entonces, el experimentador puede trazar la velocidad o velocidad de reacción inicial (Vo) de una prueba determinada frente a la concentración del sustrato ([S]) para obtener una gráfica de las propiedades de la reacción. Para muchas reacciones enzimáticas que involucran un solo sustrato, estos datos se ajustan a la ecuación de Michaelis-Menten, una ecuación derivada por Leonor Michaelis y Maud Menten.

<img alt="Eq1" src="/files/ftp_upload/12164/12164_Equation 1.png" style="text-align: center; height: 70px;" />

La ecuación estima la velocidad máxima (Vmax) y la constante de Michaelis (KM) para la enzima que se está estudiando y se basa en los siguientes supuestos:

<ol>
	<li>No hay producto presente al inicio de la reacción.</li>
	<li>La tasa de formación del complejo enzima-sustrato es igual a la tasa de disociación y descomposición en productos.</li>
	<li>La concentración de enzima es mínima en comparación con la concentración de sustrato.</li>
	<li>Sólo se miden las velocidades de reacción iniciales.</li>
	<li>La enzima está presente en forma libre o en el complejo enzima-sustrato.</li>
</ol>

Diferentes reordenamientos de la ecuación de Michaelis-Menten, como los gráficos de Lineweaver-Burke, Eadie-Hofsteot y Hanes-Woolf, son formas alternativas de representar gráficamente los parámetros cinéticos. Para estimar el KM y la Vmax se suele utilizar el gráfico de Lineweaver-Burke o doble recíproco. El gráfico utiliza los valores recíprocos de los ejes x e y del gráfico de Michaelis-Menten. Matemáticamente, la intersección con el eje y es igual a 1/Vmax y la intersección con el eje x es igual a −1/KM.

El gráfico de Lineweaver-Burke se puede utilizar para diferenciar visualmente entre tipos de inhibidores: competitivos, no competitivos y no competitivos. También se utilizan diferentes reordenamientos de la ecuación de Michaelis-Menten, como los gráficos de Eadie-Hofstee y Hanes-Woolf, para determinar los parámetros cinéticos.

Introducción a la cinética enzimática

La cinética enzimática estudia las velocidades de las reacciones bioquímicas. Los científicos monitorean las velocidades de reacción para una reacción enzimática particular ...

Education

Educación

JoVE Core

Cell Biology

Biología Celular

Unit 1

Unidad 1

Energy and Catalysis

Energía y Catálisis

KEY TERMS AND CONCEPTS

TÉRMINOS Y CONCEPTOS CLAVE

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

the first law of thermodynamics

Energy Changes in Cells and Matter

The first law of thermodynamics describes how energy changes form in the universe. The total amount of energy stays constant. Energy can move from one place to another or change into a different form, but it cannot be created or destroyed. These energy changes happen in everyday life. A light bulb changes electrical energy into light energy. A gas stove changes chemical energy in natural gas into heat energy. Plants also carry out a major energy transformation when they convert sunlight energy...

the second law of thermodynamics

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Entropy helps predict whether a process will happen on its own. Entropy is a measure of randomness or disorder in a system. High entropy means high disorder and low energy. A student’s bedroom is a simple example. If no work is done, the room becomes messy over time. That messy state is highly disordered and has high entropy. To make the room clean again, energy must be added as the student puts things away. That organized state has low entropy. Processes that increase the entropy of the...

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entropy within the cell

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the-equilibrium-binding-constant-and-binding-strength

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introduction to enzyme kinetics

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...

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introduction to mechanisms of enzyme catalysis

introduction-to-mechanisms-of-enzyme-catalysis

How Enzymes Change Shape to Catalyze

Enzyme catalysis depends on how enzymes bind substrates and how the active site responds during the reaction. Scientists once described this process with a simple lock-and-key model, where the enzyme and substrate fit together in one instant. Research now supports induced fit, a more dynamic model in which binding causes a small change in the enzyme’s shape. That shift helps create an ideal match between the enzyme and the substrate’s transition state, which improves catalysis. The active site...

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key terms and concepts

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

cell biology

JoVE Core Biología Celular ilustra procesos celulares complejos mediante completos vídeos animados fáciles de entender. Esta colección se centra en los componentes esenciales de las células y su funcionamiento, incluyendo los orgánulos y las membranas, el transporte celular y el metabolismo energético celular. Todo este material se complementa con los vídeos de científicos en acción, que muestran la aplicación de la biología celular en experimentos reales de investigación.

Virginia  Standards of Learning

HIgh School

BIO.2

BIO.2.c

Chemistry

CH.7

CH.7.f

Florida Department of Education

High School

Life Science

SC.912.L.18

SC.912.L.18.4

SC.912.L.18.11

Texas Essential Knowledge and Skills

BIO.11

BIO.11.B

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Nucleic Acid Structure

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LS.Bio.1.1

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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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Ligand Binding and Linkage

Cooperative Allosteric Transitions

Ribozymes

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Cytoplasm

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The nucleus is a membrane-bound organelle that acts as a control center in a eukaryotic cell. It contains chromosomal DNA, which controls gene expression ...

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Analyze how the relationship between structure and function supports life processes within organisms.

Graphing Enzyme Kinetics Data

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