Team:Imperial College London/Drylab/Enzyme/Analysis

From 2009.igem.org

(Difference between revisions)
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**No allosteric regulations from either the product or the substrate.
**No allosteric regulations from either the product or the substrate.
**There is no product inhibition of the enzyme.
**There is no product inhibition of the enzyme.
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*Total [E] does not change with regards to time. Enzyme will not be used up nor degraded over time, thus [E] at the beginning of the reaction will be the same as [E] at the end of reaction.
*Total [E] does not change with regards to time. Enzyme will not be used up nor degraded over time, thus [E] at the beginning of the reaction will be the same as [E] at the end of reaction.
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*For which  
*For which  
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<math> \begin{align}
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[[Image:ek1.jpg]]
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E + S
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\underset{k_{2}}{\overset{k_{1}}
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{\begin{smallmatrix}\displaystyle\longrightarrow \\ \displaystyle\longleftarrow \end{smallmatrix}}}
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ES
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\overset{k_3}
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{\longrightarrow}
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E + P
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\end{align}</math>
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, k1>>k2
, k1>>k2
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<br>
<b>Law of mass action assumptions:</b>
<b>Law of mass action assumptions:</b>
*Free diffusion
*Free diffusion
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<b>Michaelis-Menten assumption:</b>
<b>Michaelis-Menten assumption:</b>
*There is a quasi-steady state of [ES] , ie:  , during which the enzyme is used for the reaction. This means that the rate of formation of ES complex is equal to the rate of dissociation of ES complex.
*There is a quasi-steady state of [ES] , ie:  , during which the enzyme is used for the reaction. This means that the rate of formation of ES complex is equal to the rate of dissociation of ES complex.
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*<math> \begin{align}
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*
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K_M^{\prime} \ &\stackrel{\mathrm{def}}{=}\  \frac{k_3}{k_2 + k_3} K_M = \frac{k_3}{k_2 + k_3} \cdot \frac{k_{2} + k_{-1}}{k_{1}}\\
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k_{cat} \ &\stackrel{\mathrm{def}}{=}\  \dfrac{k_3 k_2}{k_2 + k_3}
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\end{align} </math>
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*from the above assumptions, we know that   
*from the above assumptions, we know that   
{{Imperial/09/TemplateBottom}}
{{Imperial/09/TemplateBottom}}

Revision as of 04:42, 22 September 2009



Assumptions

Enzymatic assumptions:

  • The enzyme is specific only for the substrate and not for any other chemicals.
  • Only one enzyme, our enzyme of interest is present and participating in the reaction.
  • There is negligible formation of product without the enzyme.
  • The rate of enzymatic activity remains constant over time because:
    • There is no co-operativity of the system. Binding of substrate to one enzyme binding site doesn't influence the affinity or activity of an adjacent site.
    • No allosteric regulations from either the product or the substrate.
    • There is no product inhibition of the enzyme.


  • Total [E] does not change with regards to time. Enzyme will not be used up nor degraded over time, thus [E] at the beginning of the reaction will be the same as [E] at the end of reaction.
  • The reaction catalysed is irreversible
  • [S] >> [E], such that the free concentration of substrate is very close to the concentration I added. This also ensures a constant substrate concentration throughout the assay. This allows easy determination of [E].
  • For which

Ek1.jpg

, k1>>k2


Law of mass action assumptions:

  • Free diffusion
  • Free unrestricted molecular motion


Michaelis-Menten assumption:

  • There is a quasi-steady state of [ES] , ie: , during which the enzyme is used for the reaction. This means that the rate of formation of ES complex is equal to the rate of dissociation of ES complex.
  • from the above assumptions, we know that


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