Team:Newcastle/Modelling/Degradation

From 2009.igem.org

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We used the degradation as another control in our Stochastic Switch construct in additon to IPTG and Xylose. While IPTG and Xylose control the expression of Hin, we designed an arabinose inducable degradation for Hin protein. Hence the width of the pulse for Hin expression can easily be adjusted by arabinose.
We used the degradation as another control in our Stochastic Switch construct in additon to IPTG and Xylose. While IPTG and Xylose control the expression of Hin, we designed an arabinose inducable degradation for Hin protein. Hence the width of the pulse for Hin expression can easily be adjusted by arabinose.
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SspB protein regulated by arabinose binds to the modified ssrA tag and targets the Hin for degradation by ClpXP system.
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SspB protein regulated by arabinose binds to the modified ''ssrA'' tag and targets the Hin for degradation by ClpXP system.
[[Image:Team_Newcastle_iGEM_2009_Degradation_Model_7.png]]
[[Image:Team_Newcastle_iGEM_2009_Degradation_Model_7.png]]
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We assumed that the species have the following inital concentrations. We took these values by running the model when there is no arabinose.
We assumed that the species have the following inital concentrations. We took these values by running the model when there is no arabinose.
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[[Image:Team_Newcastle_iGEM_2009_Degradation_Model_1.png|400px]]
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[[Image:Team_Newcastle_iGEM_2009_Degradation_Model_1.png|500px]]
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When the arabinose concentration is increased to 1000nM, Hin levels decrease sharply in 15 minutes.
When the arabinose concentration is increased to 1000nM, Hin levels decrease sharply in 15 minutes.
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[[Image:Team_Newcastle_iGEM_2009_Degradation_Model_2.png|400px]]
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[[Image:Team_Newcastle_iGEM_2009_Degradation_Model_2.png|500px]]
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The reason for this decrease is very obvious. Arabibose binds to AraR protein which normally repress sppB expression and sspB exprssion increases. As a result Hin proteins are targeted for degradation.
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The reason for this decrease is very obvious. Arabinose binds to AraR protein which normally repress SppB expression and SspB expression increases. As a result Hin proteins are targeted for degradation.
-
[[Image:Team_Newcastle_iGEM_2009_Degradation_Model_3.png|400px]]
+
[[Image:Team_Newcastle_iGEM_2009_Degradation_Model_3.png|500px]]
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We haven't gone into much detail about the rate parameters of the ClpXP degradation system. We assumed that the system would degrade a 0.2 protein per second. However we changed this parameter between 0.1 and 0,8 at 100 intervals. Range of sspB and Hin concentrations are displayed below against 100 different values of the protein degradation speed of ClpXP system.
+
We haven't gone into much detail about the rate parameters of the ClpXP degradation system. We assumed that the system would degrade a 0.2 protein per second. However we changed this parameter between 0.1 and 0,8 at 100 intervals. Range of SspB and Hin concentrations are displayed below against 100 different values of the protein degradation speed of ClpXP system.
-
[[Image:Team_Newcastle_iGEM_2009_Degradation_Model_4.png|400px]]
+
[[Image:Team_Newcastle_iGEM_2009_Degradation_Model_4.png|500px]]
The diagran below shows the concentration of total Hin proteins (proteins that are free and bound to sspB) against SspB protein. X-axis represents SspB and Y-axis represents Hin. As it can be seen when SspB reached to its maximum level, Hin has its the lowest values,
The diagran below shows the concentration of total Hin proteins (proteins that are free and bound to sspB) against SspB protein. X-axis represents SspB and Y-axis represents Hin. As it can be seen when SspB reached to its maximum level, Hin has its the lowest values,
-
[[Image:Team_Newcastle_iGEM_2009_Degradation_Model_5.png|400px]]
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[[Image:Team_Newcastle_iGEM_2009_Degradation_Model_5.png|500px]]
We also wanted to investigate how different levels of arabinose effect the system. In the diagram below each line represents Hin levels against different arabinose concentration. Arabinose levels are changed between 0nM and 10000nM with 20 intervals with 500nM interval step. The top line is when the arabinose concentration is 0nM and the bottom line is when arabinose concenration 10000nM. In the later case, Hin proteins are degraded in 7-8 minutes.
We also wanted to investigate how different levels of arabinose effect the system. In the diagram below each line represents Hin levels against different arabinose concentration. Arabinose levels are changed between 0nM and 10000nM with 20 intervals with 500nM interval step. The top line is when the arabinose concentration is 0nM and the bottom line is when arabinose concenration 10000nM. In the later case, Hin proteins are degraded in 7-8 minutes.
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[[Image:Team_Newcastle_iGEM_2009_Degradation_Model_6.png|400px]]
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[[Image:Team_Newcastle_iGEM_2009_Degradation_Model_6.png|500px]]
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Finally this animation shows the increase and decrease in each species' concentration. The volume of the balls corerspond to their concentrations.
Finally this animation shows the increase and decrease in each species' concentration. The volume of the balls corerspond to their concentrations.
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[[Image:Team_Newcastle_iGEM_2009_Degradation_Model_8.gif|400px]]
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[[Image:Team_Newcastle_iGEM_2009_Degradation_Model_8.gif|500px]]
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*Modifiers are listed after '';'' character
*Modifiers are listed after '';'' character
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[[Image:Team_Newcastle_iGEM_2009_Degradation_Model_9.png|400px]]
 
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[[Image:Team_Newcastle_iGEM_2009_Degradation_Model_9.png]]
===ODES===
===ODES===
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[[Image:Team_Newcastle_iGEM_2009_Degradation_Model_10.png|400px]]
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[[Image:Team_Newcastle_iGEM_2009_Degradation_Model_10.png|600px]]
===Model===
===Model===
Model can be downloaded in SBML format.
Model can be downloaded in SBML format.
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[[Media:Team_Newcastle_iGEM_2009_DegradationControl_SBML_Model.zip‎ SBML Model]]
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[https://static.igem.org/mediawiki/2009/e/e4/Team_Newcastle_iGEM_2009_DegradationControl_SBML_Model.zip SBML Model]
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{{:Team:Newcastle/Footer}}
{{:Team:Newcastle/Footer}}
{{:Team:Newcastle/Right}}
{{:Team:Newcastle/Right}}

Latest revision as of 23:39, 21 October 2009

Contents

Degradation Model

We used the degradation as another control in our Stochastic Switch construct in additon to IPTG and Xylose. While IPTG and Xylose control the expression of Hin, we designed an arabinose inducable degradation for Hin protein. Hence the width of the pulse for Hin expression can easily be adjusted by arabinose.

SspB protein regulated by arabinose binds to the modified ssrA tag and targets the Hin for degradation by ClpXP system.

Team Newcastle iGEM 2009 Degradation Model 7.png

We assumed that the species have the following inital concentrations. We took these values by running the model when there is no arabinose.

Team Newcastle iGEM 2009 Degradation Model 1.png


When the arabinose concentration is increased to 1000nM, Hin levels decrease sharply in 15 minutes.

Team Newcastle iGEM 2009 Degradation Model 2.png


The reason for this decrease is very obvious. Arabinose binds to AraR protein which normally repress SppB expression and SspB expression increases. As a result Hin proteins are targeted for degradation.

Team Newcastle iGEM 2009 Degradation Model 3.png


We haven't gone into much detail about the rate parameters of the ClpXP degradation system. We assumed that the system would degrade a 0.2 protein per second. However we changed this parameter between 0.1 and 0,8 at 100 intervals. Range of SspB and Hin concentrations are displayed below against 100 different values of the protein degradation speed of ClpXP system.

Team Newcastle iGEM 2009 Degradation Model 4.png


The diagran below shows the concentration of total Hin proteins (proteins that are free and bound to sspB) against SspB protein. X-axis represents SspB and Y-axis represents Hin. As it can be seen when SspB reached to its maximum level, Hin has its the lowest values,

Team Newcastle iGEM 2009 Degradation Model 5.png


We also wanted to investigate how different levels of arabinose effect the system. In the diagram below each line represents Hin levels against different arabinose concentration. Arabinose levels are changed between 0nM and 10000nM with 20 intervals with 500nM interval step. The top line is when the arabinose concentration is 0nM and the bottom line is when arabinose concenration 10000nM. In the later case, Hin proteins are degraded in 7-8 minutes.

Team Newcastle iGEM 2009 Degradation Model 6.png


Finally this animation shows the increase and decrease in each species' concentration. The volume of the balls corerspond to their concentrations.

Team Newcastle iGEM 2009 Degradation Model 8.gif


Reactions

We used COPASI to prepare this model. The tool has its own format to represent the recations.

  • ->:Irreversible reactions
  • = :Reversible reactions
  • Modifiers are listed after ; character


Team Newcastle iGEM 2009 Degradation Model 9.png

ODES

Team Newcastle iGEM 2009 Degradation Model 10.png

Model

Model can be downloaded in SBML format.

SBML Model




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