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

Kinetyka enzymów bada szybkość reakcji katalizowanych enzymami. Tempo powtarzania eksperymentów przy różnych stężeniach substratów jest monitorowane poprzez pomiar stężenia zużytego substratu lub produktu powstałego w czasie.
Wyniki te można przedstawić na wykresie, aby pokazać, w jaki sposób stężenie substratu wpływa na szybkość lub szybkość reakcji.
Szybkość reakcji wzrasta liniowo wraz ze wzrostem ilości substratu przy niskich stężeniach, ale zaczyna się stabilizować przy wyższych stężeniach. Szybkość zbliża się do maksymalnej prędkości lub Vmax – szybkości, w której enzym jest całkowicie nasycony substratem.
Powinowactwo enzymatyczne mierzy, jak silnie lub słabo enzym wiąże swój substrat i jest określane ilościowo przez KM, stałą Michaelisa. Wartość KM jest równa stężeniu substratu, gdy szybkość wynosi 50% Vmax.
Małe KM oznacza, że enzym ma wysokie powinowactwo do substratu i odwrotnie. Enzym o większym KM wymaga wyższych stężeń substratu, aby zbliżyć się do maksymalnej prędkości w porównaniu z enzymem o niższym KM.

<ol>
	<li>Voet D. i Voet J.G. Biochemia, 4 wyd. John Wiley & Sons, 2010, s. 487-492</li>
	<li>Berg J.M., Tymoczko J.L. i Stryer L. Biochemia, wyd. 5, WH Freeman and Co., sekcja 8.4</li>
</ol>

wprowadzenie do kinetyki enzymów

Kinetyka enzymów bada szybkość reakcji biochemicznych. Naukowcy monitorują szybkość reakcji określonej reakcji enzymatycznej przy różnych stężeniach substratu. Można również przeprowadzić dodatkowe próby z inhibitorami lub innymi cząsteczkami wpływającymi na szybkość reakcji. Eksperymentator może następnie wykreślić początkową szybkość lub prędkość reakcji (Vo) w danej próbie w funkcji stężenia substratu ([S]), aby uzyskać wykres właściwości reakcji. W przypadku wielu reakcji enzymatycznych obejmujących pojedynczy substrat dane te pasują do równania Michaelisa-Mentena, równania wyprowadzonego przez Leonor Michaelis i Maud Menten. <img alt="Eq1" src="/files/ftp_upload/12164/12164_Equation 1.png" style="text-align: center; height: 70px;"> Równanie szacuje maksymalną prędkość (Vmax) i stałą Michaelisa (KM) dla badanego enzymu i opiera się na następujących założeniach: <ol> <li>Na początku reakcji nie ma żadnego produktu.</li> <li>Szybkość tworzenia kompleksu enzym-substrat jest równa szybkości dysocjacji i rozkładu na produkty.</li> <li>Stężenie enzymu jest minimalne w porównaniu do stężenia substratu.</li> <li>Mierzy się jedynie początkowe szybkości reakcji.</li> <li>Enzym występuje w postaci wolnej lub w kompleksie enzym-substrat.</li> </ol> Różne przegrupowania równania Michaelisa-Menten, takie jak wykresy Lineweavera-Burka, Eadie-Hofstee i Hanesa-Woolfa, to alternatywne sposoby wykreślania parametrów kinetycznych. Do oszacowania KM i Vmax często stosuje się wykres Lineweavera-Burka lub wykres podwójnej odwrotności. Na wykresie wykorzystano odwrotności wartości osi x i y z wykresu Michaelisa-Menten. Matematycznie, punkt przecięcia y wynosi 1/Vmax, a punkt przecięcia x wynosi −1/KM. Wykres Lineweavera-Burka można wykorzystać do wizualnego rozróżnienia typów inhibitorów - konkurencyjnych, niekonkurencyjnych i niekonkurencyjnych. Do określenia parametrów kinetycznych stosuje się również różne przegrupowania równania Michaelisa-Menten, takie jak wykresy Eadie-Hofstee i Hanes-Woolf.

Wprowadzenie do kinetyki enzymów

Kinetyka enzymów bada szybkość reakcji biochemicznych. Naukowcy monitorują szybkość reakcji dla określonej reakcji enzymatycznej na różnych podłożach ...

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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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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

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Virginia  Standards of Learning

HIgh School

BIO.2

BIO.2.c

Chemistry

CH.7

CH.7.f

Florida Department of Education

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Life Science

SC.912.L.18

SC.912.L.18.4

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BIO.11

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Graphing Enzyme Kinetics Data

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