Difference between revisions of "Team:NCKU Tainan/Model"
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<body data-spy="scroll" data-target=".navbar-example"> | <body data-spy="scroll" data-target=".navbar-example"> | ||
<div class="container content"> | <div class="container content"> | ||
| − | + | <div class="headstyle"> | |
| + | <h1 class="head">Model</h1> | ||
| + | </div> | ||
| + | <div class="righttitle"> | ||
| + | <h6 class="subtitle">Prediction of Metabolism</h6> | ||
| + | </div> | ||
<div class="navbar-example"> | <div class="navbar-example"> | ||
<div class="row"> | <div class="row"> | ||
| − | <div class="col-2 side"> | + | <div class="col-2 side"> |
<div id="sidelist" class="list-group"> | <div id="sidelist" class="list-group"> | ||
<a class="list-group-item list-group-item-action" href="#achievement">Achievement</a> | <a class="list-group-item list-group-item-action" href="#achievement">Achievement</a> | ||
<a class="list-group-item list-group-item-action" href="#purpose">Purpose</a> | <a class="list-group-item list-group-item-action" href="#purpose">Purpose</a> | ||
| + | <a class="list-group-item list-group-item-action" href="#method">Method</a> | ||
<a class="list-group-item list-group-item-action" href="#CO2_uptake">CO<sub>2</sub> uptake</a> | <a class="list-group-item list-group-item-action" href="#CO2_uptake">CO<sub>2</sub> uptake</a> | ||
<a class="list-group-item list-group-item-action" href="#CO2_metabolism">CO<sub>2</sub> metabolism</a> | <a class="list-group-item list-group-item-action" href="#CO2_metabolism">CO<sub>2</sub> metabolism</a> | ||
<a class="list-group-item list-group-item-action" href="#CO2_vs_xylose">Analysis</a> | <a class="list-group-item list-group-item-action" href="#CO2_vs_xylose">Analysis</a> | ||
| − | <a class="list-group-item list-group-item-action" href="# | + | <a class="list-group-item list-group-item-action" href="#references">References</a> |
<a class="list-group-item list-group-item-action" href="#"><i class="fa fa-arrow-up fa-1x" aria-hidden="true"></i></a> | <a class="list-group-item list-group-item-action" href="#"><i class="fa fa-arrow-up fa-1x" aria-hidden="true"></i></a> | ||
</div> | </div> | ||
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<div id="achievement"> | <div id="achievement"> | ||
<h3>Achievement</h3> | <h3>Achievement</h3> | ||
| − | <p></p> | + | <div class="achievementborder"> |
| + | <ol> | ||
| + | <br> | ||
| + | <li class="bigli">Figure out how much CO<sub>2</sub> uptake into the <i>E.coli</i> to make our project more effective.</li> | ||
| + | <br> | ||
| + | <li class="bigli">We will realize how much carbon we can fix in our project.</li> | ||
| + | <br> | ||
| + | </ol> | ||
| + | </div> | ||
| + | <img class="gif" src="https://static.igem.org/mediawiki/2018/b/bb/T--NCKU_Tainan--model_model.gif" alt="Model"> | ||
| + | <br> | ||
| + | <p class="pcontent">In order to achieve these two goals, we try to figure out three main questions：</p> | ||
| + | <ol> | ||
| + | <li class="licontent"><a class="link" href="#CO2_uptake">Amount of CO<sub>2</sub> uptake by <i>E. coli</i></a></li> | ||
| + | <li class="licontent"><a class="link" href="https://2018.igem.org/Team:NCKU_Tainan/Kinetic_Law">CO<sub>2</sub> flux in <i>E. coli</i> metabolism</a></li> | ||
| + | <li class="licontent"><a class="link" href="https://2018.igem.org/Team:NCKU_Tainan/Analysis">Amount of Carbon fixed in biomass</a></li> | ||
| + | </ol> | ||
</div> | </div> | ||
<div id="purpose"> | <div id="purpose"> | ||
<h3>Purpose</h3> | <h3>Purpose</h3> | ||
| − | <p></p> | + | <p class="pcontent">We aim to develop the biological model to make our experimental data more useful, |
| + | even more effective to achieve the optimization parameters in the model. | ||
| + | Therefore, we can save time to try and error on doing experiment. | ||
| + | After that, we analyze the rate of production and consumption. | ||
| + | In this way, we can calculate the amount of CO<sub>2</sub> uptake into the | ||
| + | <i>Escherichia coli </i> ( <i>E. coli</i> ) and | ||
| + | calculate how much CO<sub>2</sub> will be used in our system. | ||
| + | In addition, we also want to realize how much carbon being fixed in our system. | ||
| + | We have to understand the process in <i>E. coli</i> after uptaking | ||
| + | CO<sub>2</sub>. | ||
| + | Since we integrated non-native carbon fixation pathway into <i>E. coli</i> to let | ||
| + | <i>E. coli</i> utilize CO<sub>2</sub>, | ||
| + | we simplify the CO<sub>2</sub> utilization pathway in engineered <i>E. coli</i> | ||
| + | into two parts, | ||
| + | <a class="link" href="#CO2_uptake">CO<sub>2</sub> uptake</a> and <a class="link" href="#CO2_metabolism">CO<sub>2</sub> metabolism</a>. | ||
| + | </p> | ||
| + | </div> | ||
| + | <div id="method"> | ||
| + | <h3>Method of model</h3> | ||
| + | <p class="pcontent">Follow our model progress diagram with a better sense of how we achieve our goal.</p> | ||
| + | <div class="row"> | ||
| + | <div class="col-2"></div> | ||
| + | <div class="col-8" id="cube1"> | ||
| + | <h4 class="pb">Purpose of model</h4> | ||
| + | <p class="pb">How many CO<sub>2</sub> can be used by <i>E. coli</i></p> | ||
| + | </div> | ||
| + | <div class="col-2"></div> | ||
| + | </div> | ||
| + | <div class="row"> | ||
| + | <div class="col-12" id="methodicon"> | ||
| + | <p class="pcontent" id="arrowdown"><i class="fa fa-arrow-down"></i></p> | ||
| + | </div> | ||
| + | </div> | ||
| + | <div class="row"> | ||
| + | <div class="col-2"></div> | ||
| + | <div class="col-8" id="cube2"> | ||
| + | <a class="methodlink" href="https://2018.igem.org/Team:NCKU_Tainan/Kinetic_Law"> | ||
| + | <h4 class="pb">Method of model</h4> | ||
| + | <p class="pb">Kinetic law / Metabolic Pathway / Chemical Equation</p> | ||
| + | </a> | ||
| + | </div> | ||
| + | <div class="col-2"></div> | ||
| + | </div> | ||
| + | <div class="row"> | ||
| + | <div class="col-12" id="methodicon"> | ||
| + | <p class="pcontent" id="arrowdown"><i class="fa fa-arrow-down"></i></p> | ||
| + | </div> | ||
| + | </div> | ||
| + | <div class="row"> | ||
| + | <div class="col-2"></div> | ||
| + | <div class="col-8" id="cube1"> | ||
| + | <a class="methodlink" href="https://2018.igem.org/Team:NCKU_Tainan/Analysis"> | ||
| + | <h4 class="pb">Collect experiment data</h4> | ||
| + | </a> | ||
| + | </div> | ||
| + | <div class="col-2"></div> | ||
| + | </div> | ||
| + | <div class="row"> | ||
| + | <div class="col-12" id="methodicon"> | ||
| + | <p class="pcontent" id="arrowdown"><i class="fa fa-arrow-down"></i></p> | ||
| + | </div> | ||
| + | </div> | ||
| + | <div class="row"> | ||
| + | <div class="col-2"></div> | ||
| + | <div class="col-8" id="cube3"> | ||
| + | <a class="methodlink" href="https://2018.igem.org/Team:NCKU_Tainan/Analysis"> | ||
| + | <h4 class="pb">Result analysis</h4> | ||
| + | </a> | ||
| + | </div> | ||
| + | <div class="col-2"></div> | ||
| + | </div> | ||
</div> | </div> | ||
<div id="CO2_uptake"> | <div id="CO2_uptake"> | ||
<h3>CO<sub>2</sub> uptake</h3> | <h3>CO<sub>2</sub> uptake</h3> | ||
| − | <p></p> | + | <p class="pcontent">CO<sub>2</sub> exists in the phase of gas. |
| + | Therefore, <i>E. coli</i> uptake CO<sub>2</sub> by diffusion. | ||
| + | However, the native diffusion rate is really low, not to mention the dissolved rate. | ||
| + | We clone CA (carbonic anhydrase) gene to speed up the diffusion rate. | ||
| + | The higher diffusion rate, the higher CO<sub>2</sub> concentration maintained in engineered <i>E. coli</i>. | ||
| + | Higher CO<sub>2</sub> concentration in engineered <i>E. coli</i> provides more steady CO<sub>2</sub> condition for downstream pathway.</p> | ||
<div class="row"> | <div class="row"> | ||
<img class="col-md-12 smallimg" src="https://static.igem.org/mediawiki/2018/0/06/T--NCKU_Tainan--model_uptake.png"> | <img class="col-md-12 smallimg" src="https://static.igem.org/mediawiki/2018/0/06/T--NCKU_Tainan--model_uptake.png"> | ||
| − | </div> | + | </div> |
<div> | <div> | ||
<div class="row"> | <div class="row"> | ||
<a class="btn col-md-12" data-toggle="collapse" href="#co2_uptake" role="button" aria-expanded="false" aria-controls="multiCollapseExample1"> | <a class="btn col-md-12" data-toggle="collapse" href="#co2_uptake" role="button" aria-expanded="false" aria-controls="multiCollapseExample1"> | ||
| − | CO<sub>2</sub> uptake | + | Table 1. parameters of CO<sub>2</sub> uptake pathway |
<i class="fa fa-arrow-down fa-10" aria-hidden="true"></i> | <i class="fa fa-arrow-down fa-10" aria-hidden="true"></i> | ||
</a> | </a> | ||
| − | </div> | + | </div> |
| − | + | <div class="collapse multi-collapse" id="co2_uptake"> | |
| − | + | <div class="card card-body"> | |
| − | + | <table> | |
| − | + | ||
<tr> | <tr> | ||
<th colspan="4">CO<sub>2</sub> uptake</th> | <th colspan="4">CO<sub>2</sub> uptake</th> | ||
| − | <th colspan="2"> | + | <th colspan="2">K<sub>f</sub> (mM)</th> |
| − | <th colspan="2"> | + | <th colspan="2">K<sub>r</sub> (mM)</th> |
</tr> | </tr> | ||
<tr> | <tr> | ||
<td colspan="1">R1</td> | <td colspan="1">R1</td> | ||
| − | <td colspan="3">$${[CO_2]_{air} \rightarrow [CO_2] | + | <td colspan="3">$${[CO_2]_{air} \rightarrow [CO_2]_{uptake}}$$</td> |
<td colspan="2">9.23E-04</td> | <td colspan="2">9.23E-04</td> | ||
<td colspan="2">4.78E-04</td> | <td colspan="2">4.78E-04</td> | ||
| Line 80: | Line 176: | ||
<td colspan="1" rowspan="2">R5</td> | <td colspan="1" rowspan="2">R5</td> | ||
<td colspan="3" rowspan="2">$${[H^+] + [HCO_3^-] \longleftrightarrow [H_2O] + [CO_2]_{uptake}}$$</td> | <td colspan="3" rowspan="2">$${[H^+] + [HCO_3^-] \longleftrightarrow [H_2O] + [CO_2]_{uptake}}$$</td> | ||
| − | <td colspan="2"> | + | <td colspan="2">V<sub>max</sub></td> |
| − | <td colspan="2"> | + | <td colspan="2">K<sub>m</sub></td> |
</tr> | </tr> | ||
<tr> | <tr> | ||
| Line 87: | Line 183: | ||
<td colspan="2">0.0062</td> | <td colspan="2">0.0062</td> | ||
</tr> | </tr> | ||
| − | + | </table> | |
| − | </div> | + | </div> |
| + | </div> | ||
| + | <div class="row"> | ||
| + | <h4>Result</h4> | ||
| + | <p class="pcontent">Enzyme, CA, strongly catalyzes the hydration of CO<sub>2</sub> and its interaction appears to | ||
| + | follow a Michaelis-Menten mechanism with a Hill constant of 0.0062 in our research. | ||
| + | The result of Engineered <i>E. coli</i> cloned with or without CA gene showed below. | ||
| + | A quite different diffusion time explains the function of CA. | ||
| + | </p> | ||
| + | <div class="col-12" id="resultimg"> | ||
| + | <img class="twoimg" src="https://static.igem.org/mediawiki/2018/7/75/T--NCKU_Tainan--model_result1.png"> | ||
| + | <p class="pcenter">Fig 1. CO<sub>2</sub> uptake amount change with time in engineered <i>E. coli</i> without CA gene.</p> | ||
| + | <img class="twoimg" src="https://static.igem.org/mediawiki/2018/9/9f/T--NCKU_Tainan--model_result2.png"> | ||
| + | <p class="pcenter">Fig 2. CO<sub>2</sub> uptake amount change with time in engineered <i>E. coli</i> with CA gene.</p> | ||
</div> | </div> | ||
| + | </div> | ||
</div> | </div> | ||
</div> | </div> | ||
| Line 95: | Line 205: | ||
<div id="CO2_metabolism"> | <div id="CO2_metabolism"> | ||
<h3>CO<sub>2</sub> Metabolism</h3> | <h3>CO<sub>2</sub> Metabolism</h3> | ||
| − | <div> | + | <p class="pcontent">Xylose is the main source of our engineered <i>E. coli</i> to uptake CO<sub>2</sub>. |
| + | However, there is another pathway for xylose metabolism, | ||
| + | which means that there is only a part of xylose that <i>E. coli</i> consumed will react with CO<sub>2</sub>. | ||
| + | We build a model to predict the different carbon flux between original xylose metabolism, | ||
| + | <a class="link" href="#ppptable">PP pathway (Pentose Phosphate pathway)</a>, and our <a class="link" href="#bypasstable">CO<sub>2</sub> bypass pathway</a>. | ||
| + | </p> | ||
| + | <p class="pcontent">Both of two pathways will end up at pyruvate, | ||
| + | which is a key intersection in the network of metabolic pathways. | ||
| + | We built a model of <a class="link" href="#tcatable">TCA cycle</a> to explain how pyruvate play an energy precursor role and then | ||
| + | is converted into biomass of <i>E. coli</i>. | ||
| + | </p> | ||
| + | <div class="row"> | ||
| + | <h3>Pathway</h3> | ||
| + | <div class="col-12"> | ||
| + | <img class="col-md-6" id="easypathway" src="https://static.igem.org/mediawiki/2018/f/f8/T--NCKU_Tainan--model_easypathway.png"> | ||
| + | <img class="col-md-6 contentimg" src="https://static.igem.org/mediawiki/2018/3/3e/T--NCKU_Tainan--model_pathway.png"> | ||
| + | <p class="pcenter col-12">Fig 3. Metabolic pathway of CO<sub>2</sub>-utilization <i>E. coli</i>. </p> | ||
| + | <p class="pcontent col-12">The introduced CO<sub>2</sub>-utilization bypass pathway composed of PRK and Rubisco is drawn in green line and noted by ” A” reaction and the double line was the pathway that composed by genetically modified, while the central carbon metabolic pathway including PP pathway and TCA cycle is drawn in blue line and yellow line and noted by “B” reaction and “C” reaction, respectively. | ||
| + | See more detail in <a class="link" href="https://2018.igem.org/Team:NCKU_Tainan/Kinetic_Law">Kinetic law</a>. | ||
| + | </p> | ||
| + | |||
| + | </div> | ||
| + | </div> | ||
| + | <div class="row"> | ||
| + | <div class="table" id="bypasstable"> | ||
| + | <div class="row"> | ||
| + | <a class="btn col-md-12" data-toggle="collapse" href="#co2_bypass" role="button" aria-expanded="false" aria-controls="multiCollapseExample1"> | ||
| + | Table 2. Kinetic parameters of CO<sub>2</sub>-utilization bypass pathway | ||
| + | <i class="fa fa-arrow-down fa-10" aria-hidden="true"></i> | ||
| + | </a> | ||
| + | </div> | ||
| + | <div class="collapse multi-collapse" id="co2_bypass"> | ||
| + | <div class="card card-body"> | ||
| + | <table> | ||
| + | <tr> | ||
| + | <th colspan="8">CO<sub>2</sub> bypass pathway</th> | ||
| + | <th colspan="3">V<sub>max</sub> (1/S)</th> | ||
| + | <th colspan="3">K<sub>M</sub> (mM)</th> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="2">A1</td> | ||
| + | <td colspan="6">$${Xylose \rightarrow Xylulose}$$ | ||
| + | $${Xylulose + ATP \rightarrow X5P}$$ | ||
| + | </td> | ||
| + | <td colspan="3">0.02456</td> | ||
| + | <td colspan="3">124.55879</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="2">A2</td> | ||
| + | <td colspan="6">$${X5P \rightarrow Ru5P}$$</td> | ||
| + | <td colspan="3">0.08</td> | ||
| + | <td colspan="3">12.5</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="2">A3</td> | ||
| + | <td colspan="6">$${Ru5P \xrightarrow{PRK} RuBP}$$</td> | ||
| + | <td colspan="3">225.81858</td> | ||
| + | <td colspan="3">0.27</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td rowspan="2" colspan="2">A4</td> | ||
| + | <td rowspan="2" colspan="6">$${RuBP + CO_2 \xrightarrow{RuBisCO} 3PG}$$</td> | ||
| + | <td colspan="2">V<sub>max</sub></td> | ||
| + | <td colspan="2">K<sub>rubp</sub></td> | ||
| + | <td colspan="2">K<sub>CO</sub><sub>2</sub></td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="2">333.33</td> | ||
| + | <td colspan="2">53</td> | ||
| + | <td colspan="2">2.7</td> | ||
| + | </tr> | ||
| + | </table> | ||
| + | </div> | ||
| + | </div> | ||
| + | </div> | ||
| + | |||
| + | <div class="table" id="ppptable"> | ||
| + | <div class="row"> | ||
| + | <a class="btn col-md-12" data-toggle="collapse" href="#ppp" role="button" aria-expanded="false" aria-controls="multiCollapseExample1"> | ||
| + | Table 3. Kinetic parameters of PP pathway in <i>E. coli</i> | ||
| + | <i class="fa fa-arrow-down fa-10" aria-hidden="true"></i> | ||
| + | </a> | ||
| + | </div> | ||
| + | <div class="collapse multi-collapse" id="ppp"> | ||
| + | <div class="card card-body"> | ||
| + | <table> | ||
| + | <tr> | ||
| + | <th colspan="13">Pentose Phosphate Pathway</th> | ||
| + | <th colspan="6">V<sub>max</sub> (1/S)</th> | ||
| + | <th colspan="6">K<sub>M</sub> (mM)</th> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="1">B1</td> | ||
| + | <td colspan="12">$${Ru5P \rightarrow R5P}$$</td> | ||
| + | <td colspan="6">0.13672</td> | ||
| + | <td colspan="6">12.5</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td rowspan="5" colspan="1">B2</td> | ||
| + | <td colspan="24">$${X5P + R5P \rightarrow S7P + GAP}$$</td> </tr> | ||
| + | <tr> | ||
| + | <td colspan="12">$${X5P \rightarrow S7P}$$</td> | ||
| + | <td colspan="6">58.27</td> | ||
| + | <td colspan="6">4.96E-09</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="12">$${X5P \rightarrow GAP}$$</td> | ||
| + | <td colspan="6">58.27</td> | ||
| + | <td colspan="6">48</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="12">$${R5P \rightarrow S7P}$$</td> | ||
| + | <td colspan="6">58.27</td> | ||
| + | <td colspan="6">0.008</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="12">$${R5P \rightarrow GAP}$$</td> | ||
| + | <td colspan="6">58.27</td> | ||
| + | <td colspan="6">3.05E-07</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td rowspan="5" colspan="1">B3</td> | ||
| + | <td colspan="24">$${S7P + GAP \rightarrow F6P + E4P}$$</td> </tr> | ||
| + | <tr> | ||
| + | <td colspan="12">$${S7P \rightarrow E4P}$$</td> | ||
| + | <td colspan="6">58.27</td> | ||
| + | <td colspan="6">2.524</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="12">$${S7P \rightarrow F6P}$$</td> | ||
| + | <td colspan="6">58.27</td> | ||
| + | <td colspan="6">6.10E-09</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="12">$${GAP \rightarrow E4P}$$</td> | ||
| + | <td colspan="6">58.27</td> | ||
| + | <td colspan="6">1.73E-07</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="12">$${GAP \rightarrow F6P}$$</td> | ||
| + | <td colspan="6">58.27</td> | ||
| + | <td colspan="6">8.23E-09</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td rowspan="5" colspan="1">B4</td> | ||
| + | <td colspan="24">$${E4P + X5P \rightarrow F6P + GAP}$$</td> </tr> | ||
| + | <tr> | ||
| + | <td colspan="12">$${E4P \rightarrow F6P}$$</td> | ||
| + | <td colspan="6">1</td> | ||
| + | <td colspan="6">1</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="12">$${E4P \rightarrow GAP}$$</td> | ||
| + | <td colspan="6">5.827E-4</td> | ||
| + | <td colspan="6">1.733E-7</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="12">$${X5P \rightarrow F6P}$$</td> | ||
| + | <td colspan="6">0.05827</td> | ||
| + | <td colspan="6">1.733E-7</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="12">$${X5P \rightarrow GAP}$$</td> | ||
| + | <td colspan="6">0.005827</td> | ||
| + | <td colspan="6">48.8</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="1">B5</td> | ||
| + | <td colspan="12">$${FBP \rightarrow GAP}$$</td> | ||
| + | <td colspan="6">135.425</td> | ||
| + | <td colspan="6">4</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td rowspan="2" colspan="1">B6</td> | ||
| + | <td rowspan="2" colspan="12">$${DHAP \longleftrightarrow GAP}$$</td> | ||
| + | <td colspan="3">V<sub>mf</sub></td> | ||
| + | <td colspan="3">V<sub>mr</sub></td> | ||
| + | <td colspan="3">K<sub>m</sub><sub>DHAP</sub></td> | ||
| + | <td colspan="3">K<sub>m</sub><sub>GAP</sub></td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="3">0.2394</td> | ||
| + | <td colspan="3">0.24</td> | ||
| + | <td colspan="3">0.812</td> | ||
| + | <td colspan="3">0.245</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="1">B7</td> | ||
| + | <td colspan="12">$${FBP \rightarrow DHAP}$$</td> | ||
| + | <td colspan="6">0.0001247</td> | ||
| + | <td colspan="6">0.04</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="1">B8</td> | ||
| + | <td colspan="12">$${F6P + ATP \rightarrow FBP + ADP}$$</td> | ||
| + | <td colspan="6">88.2533</td> | ||
| + | <td colspan="6">0.017</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="1">B9</td> | ||
| + | <td colspan="12">$${F6P \rightarrow G6P}$$</td> | ||
| + | <td colspan="6">88.2353</td> | ||
| + | <td colspan="6">0.017</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td rowspan="2" colspan="1">B10</td> | ||
| + | <td rowspan="2" colspan="12">$${G6P + NADP^+ \rightarrow 6PG + NADPH}$$</td> | ||
| + | <td colspan="4">V<sub>max</sub></td> | ||
| + | <td colspan="4">K<sub>m</sub><sub>G6P</sub></td> | ||
| + | <td colspan="4">K<sub>m</sub><sub>NADP</sub></td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="4">11.736</td> | ||
| + | <td colspan="4">0.12</td> | ||
| + | <td colspan="4">0.0123</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="1">B11</td> | ||
| + | <td colspan="12">$${6PG + NADP^+ \rightarrow Ru5P + NADPH}$$</td> | ||
| + | <td colspan="6">1.0</td> | ||
| + | <td colspan="6">0.044</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="1">B12</td> | ||
| + | <td colspan="12">$${GAP + NADP^+ + ADP \rightarrow 3PGA + NADPH + ATP}$$</td> | ||
| + | <td colspan="6">0.4</td> | ||
| + | <td colspan="6">0.013</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="1">B13</td> | ||
| + | <td colspan="12">$${3PGA \rightarrow PEP}$$</td> | ||
| + | <td colspan="6">0.43478</td> | ||
| + | <td colspan="6">0.23</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="1">B14</td> | ||
| + | <td colspan="12">$${PEP \rightarrow PYR}$$</td> | ||
| + | <td colspan="6">0.23148</td> | ||
| + | <td colspan="6">0.45</td> | ||
| + | </tr> | ||
| + | </table> | ||
| + | </div> | ||
| + | </div> | ||
| + | </div> | ||
| + | |||
| + | <div class="table" id="tcatable"> | ||
| + | <div class="row"> | ||
| + | <a class="btn col-md-12" data-toggle="collapse" href="#tca" role="button" aria-expanded="false" aria-controls="multiCollapseExample1"> | ||
| + | Table 4. Kinetic parameters of TCA cycle in <i>E. coli</i> | ||
| + | <i class="fa fa-arrow-down fa-10" aria-hidden="true"></i> | ||
| + | </a> | ||
| + | </div> | ||
| + | <div class="collapse multi-collapse" id="tca"> | ||
| + | <div class="card card-body"> | ||
| + | <table> | ||
| + | <tr> | ||
| + | <th colspan="13">TCA cycle</th> | ||
| + | <th colspan="3">V<sub>mf</sub> (mM/min)</th> | ||
| + | <th colspan="3">V<sub>mr</sub> (mM/min)</th> | ||
| + | <th colspan="3">K<sub>s</sub> (mM)</th> | ||
| + | <th colspan="3">K<sub>p</sub> (mM)</th> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="1">C1</td> | ||
| + | <td colspan="12">$${PEP + HCO_3^- + NAD^+ \rightarrow OAA + NADH}$$</td> | ||
| + | <td colspan="3">1</td> | ||
| + | <td colspan="3">1</td> | ||
| + | <td colspan="3">1</td> | ||
| + | <td colspan="3">1</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="1">C2</td> | ||
| + | <td colspan="12">$${AcCoA \rightarrow CIT}$$</td> | ||
| + | <td colspan="3">64.8</td> | ||
| + | <td colspan="3">0.648</td> | ||
| + | <td colspan="3">0.05</td> | ||
| + | <td colspan="3">0.12</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="1">C3</td> | ||
| + | <td colspan="12">$${CIT \rightarrow ICIT}$$</td> | ||
| + | <td colspan="3">31.2</td> | ||
| + | <td colspan="3">0.312</td> | ||
| + | <td colspan="3">1.7</td> | ||
| + | <td colspan="3">0.7</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="1">C4</td> | ||
| + | <td colspan="12">$${ICIT + NADP^+ \rightarrow AKG + NADPH + CO_2}$$</td> | ||
| + | <td colspan="3">10.2</td> | ||
| + | <td colspan="3">0.102</td> | ||
| + | <td colspan="3">0.03</td> | ||
| + | <td colspan="3">0.3</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="1">C5</td> | ||
| + | <td colspan="12">$${AKG + NAD^+ \rightarrow AKG + NADPH + CO_2}$$</td> | ||
| + | <td colspan="3">57.344</td> | ||
| + | <td colspan="3">0.57344</td> | ||
| + | <td colspan="3">0.1</td> | ||
| + | <td colspan="3">1.0</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="1">C6</td> | ||
| + | <td colspan="12">$${SUCCoA + ADP \rightarrow SUCC + ATP}$$</td> | ||
| + | <td colspan="3">6.54</td> | ||
| + | <td colspan="3">0.0651</td> | ||
| + | <td colspan="3">0.015</td> | ||
| + | <td colspan="3">0.15</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="1">C7</td> | ||
| + | <td colspan="12">$${SUCC + FAD \rightarrow FUM + FADH_2}$$</td> | ||
| + | <td colspan="3">1.02</td> | ||
| + | <td colspan="3">1.02</td> | ||
| + | <td colspan="3">0.12</td> | ||
| + | <td colspan="3">0.15</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="1">C8</td> | ||
| + | <td colspan="12">$${FUM \rightarrow MAL}$$</td> | ||
| + | <td colspan="3">887.7</td> | ||
| + | <td colspan="3">87.7</td> | ||
| + | <td colspan="3">0.25</td> | ||
| + | <td colspan="3">2.38</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="1">C9</td> | ||
| + | <td colspan="12">$${MAL + NAD^+ \rightarrow OAA + NADH}$$</td> | ||
| + | <td colspan="3">184</td> | ||
| + | <td colspan="3">184</td> | ||
| + | <td colspan="3">0.833</td> | ||
| + | <td colspan="3">0.0443</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="1">C10</td> | ||
| + | <td colspan="12">$${OAA \rightarrow CIT}$$</td> | ||
| + | <td colspan="3">64.8</td> | ||
| + | <td colspan="3">0.648</td> | ||
| + | <td colspan="3">0.012</td> | ||
| + | <td colspan="3">0.12</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td rowspan="2" colspan="1">C11</td> | ||
| + | <td rowspan="2" colspan="12">$${PYR \rightarrow NADH + AcCoA + CO_2}$$</td> | ||
| + | <td colspan="6">V<sub>max</sub> (1/s)</td> | ||
| + | <td colspan="6">K<sub>m</sub> (mM)</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="6">0.00518</td> | ||
| + | <td colspan="6">1.5</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="1">C12</td> | ||
| + | <td colspan="12">$${ICIT \rightarrow GOX}$$</td> | ||
| + | <td colspan="3">1.172</td> | ||
| + | <td colspan="3">0.01172</td> | ||
| + | <td colspan="3">0.145</td> | ||
| + | <td colspan="3">0.13</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="1">C13</td> | ||
| + | <td colspan="12">$${GOX \rightarrow MAL}$$</td> | ||
| + | <td colspan="3">20</td> | ||
| + | <td colspan="3">0.2</td> | ||
| + | <td colspan="3">0.057</td> | ||
| + | <td colspan="3">1.0</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td rowspan="2" colspan="1">C14</td> | ||
| + | <td rowspan="2" colspan="12">$${ICIT \rightarrow SUCC + GOX}$$</td> | ||
| + | <td colspan="3">V<sub>mf</sub> (mM/min)</td> | ||
| + | <td colspan="3">V<sub>mr</sub> (mM/min)</td> | ||
| + | <td colspan="2">K<sub>icit</sub> (mM)</td> | ||
| + | <td colspan="2">K<sub>succ</sub> (mM)</td> | ||
| + | <td colspan="2">K<sub>gox</sub> (mM)</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td colspan="3">1.172</td> | ||
| + | <td colspan="3">0.012</td> | ||
| + | <td colspan="2">0.145</td> | ||
| + | <td colspan="2">0.59</td> | ||
| + | <td colspan="2">0.13</td> | ||
| + | </tr> | ||
| + | </table> | ||
| + | </div> | ||
| + | </div> | ||
| + | </div> | ||
| + | |||
<div class="table"> | <div class="table"> | ||
<div class="row"> | <div class="row"> | ||
<a class="btn col-md-12" data-toggle="collapse" href="#species" role="button" aria-expanded="false" aria-controls="multiCollapseExample1"> | <a class="btn col-md-12" data-toggle="collapse" href="#species" role="button" aria-expanded="false" aria-controls="multiCollapseExample1"> | ||
| − | Species | + | Table 5. Species of Metabolic pathway of CO<sub>2</sub>-utilization <i>E. coli</i> |
<i class="fa fa-arrow-down fa-10" aria-hidden="true"></i> | <i class="fa fa-arrow-down fa-10" aria-hidden="true"></i> | ||
</a> | </a> | ||
| Line 105: | Line 603: | ||
<div class="collapse multi-collapse" id="species"> | <div class="collapse multi-collapse" id="species"> | ||
<div class="card card-body"> | <div class="card card-body"> | ||
| − | <table | + | <table class="bbg"> |
<tr> | <tr> | ||
<td>E4P</td> | <td>E4P</td> | ||
| Line 218: | Line 716: | ||
</div> | </div> | ||
</div> | </div> | ||
| − | </div> | + | </div> |
<div class="row"> | <div class="row"> | ||
| − | + | <div class="col-12" id="CO2_vs_xylose"> | |
| − | <div class="col-12 | + | <h3>Xylose v.s. CO<sub>2</sub> analysis in CO<sub>2</sub> utilization efficiency</h3> |
| − | + | <p class="pcontent">Two main sources of CO<sub>2</sub> metabolism in engineered <i>E. coli</i> are xylose and CO<sub>2</sub>. | |
| − | < | + | CO<sub>2</sub> utilization rate varies under different condition. |
| − | + | We use 5% CO<sub>2</sub> (about 2.6 mM) and 4 (g/l) xylose (about 26 mM) in experiment as an optimization CO<sub>2</sub> utilization rate. | |
| − | + | Constant CO<sub>2</sub> condition and limited CO<sub>2</sub> condition also effects metabolism performance. | |
| − | + | In other words, open system and close system showed different results. | |
| − | + | Therefore, adjust initial concentration of CO<sub>2</sub> and Xylose and choose open or close system to see the change of pyruvate production. | |
| − | + | </p> | |
| − | < | + | <form> |
| − | + | <div class="form-row"> | |
| − | + | <div class="col"> | |
| − | + | <label for="inputState" style="color: white">Concentration of xylose</label> | |
| − | + | <select id="inputState" name="xylose" class="form-control" onclick="photos(this);"> | |
| − | + | <option value="1">0.4 %</option> | |
| − | + | <option value="2">1 %</option> | |
| − | + | <option value="3">2 %</option> | |
| − | + | </select> | |
| − | + | </div> | |
| − | + | <div class="col"> | |
| − | + | <label for="inputState" style="color: white">Concentration of CO<sub>2</sub></label> | |
| − | + | <select id="inputState" name="co2" class="form-control" onclick="photos(this);"> | |
| − | + | <option value="1">5 %</option> | |
| − | + | <option value="2">10 %</option> | |
| − | < | + | <option value="3">15 %</option> |
| − | + | </select> | |
| − | + | </div> | |
| − | < | + | <div class="col"> |
| − | + | <label for="inputState" style="color: white">Close system / Open system</label> | |
| − | + | <select id="inputState" name="co2switch" class="form-control" onclick="photos(this);"> | |
| − | + | <option value="1">Close</option> | |
| − | + | <option value="2">Open</option> | |
| − | < | + | </select> |
| − | + | </div> | |
| − | < | + | </div> |
| − | + | </form> | |
| − | + | <div class="row"> | |
| − | < | + | <img class="col-6 contentimg smallimg" style="display: none" id="MyImage"/> |
| − | + | <img class="col-6 contentimg smallimg" style="display: none" id="SwitchImage"/> | |
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</div> | </div> | ||
| − | <div | + | <div id="references"> |
| − | < | + | <h3>References</h3> |
| − | < | + | <ol> |
| − | + | <li class="smallp">E. G. Jacqueline, P. L. Christopher, R. A. Maciek, Comprehensive analysis of glucose and xylose metabolism in <i>Escherichia coli</i> under aerobic and anaerobic conditions by <sup>13</sup>C metabolic flux analysis. Metab Eng. 2017 Jan; 39: 9–18.</li> | |
| − | + | <li class="smallp">U. Sauer, J. E. Bernhard, The PEP—pyruvate—oxaloacetate node as the switch point for carbon flux distribution in bacteria. FEMS Microbiology Reviews, Volume 29, Issue 4, 1 September 2005, Pages 765–794.</li> | |
| − | + | <li class="smallp">G. Fuyu, L. Guoxia, Z. Xiaoyun, Z. Jie, C. Zhen, L. Yin . Quantitative analysis of an engineered CO<sub>2</sub>-fixing <i>Escherichia coli</i>reveals great potential of heterotrophic CO<sub>2</sub> fixation. Gong et al. Biotechnology for Biofuels, 2015, 8:86.</li> | |
| − | + | <li class="smallp">Y. Pocker, S. Y. Ng. Joan, Plant carbonic anhydrase. Properties and carbon dioxide hydration kinetics. Biochemistry, 1973, 12 (25), pp 5127–5134.</li> | |
| − | + | </ol> | |
| − | < | + | <br class="pcontent"> |
</div> | </div> | ||
</div> | </div> | ||
| Line 618: | Line 786: | ||
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| − | $('#sidelist').css({"position": "fixed", "top": " | + | $('#sidelist').css({"position": "fixed", "top": "95px"}); |
} | } | ||
} else { | } else { | ||
| − | if ($(this).scrollTop() >= | + | if ($(this).scrollTop() >= 500) { |
var position = $("#sidelist").position(); | var position = $("#sidelist").position(); | ||
if(position == undefined){} | if(position == undefined){} | ||
Latest revision as of 01:25, 18 October 2018
Model
Prediction of Metabolism
Achievement
- Figure out how much CO2 uptake into the E.coli to make our project more effective.
- We will realize how much carbon we can fix in our project.
In order to achieve these two goals, we try to figure out three main questions：
Purpose
We aim to develop the biological model to make our experimental data more useful, even more effective to achieve the optimization parameters in the model. Therefore, we can save time to try and error on doing experiment. After that, we analyze the rate of production and consumption. In this way, we can calculate the amount of CO2 uptake into the Escherichia coli ( E. coli ) and calculate how much CO2 will be used in our system. In addition, we also want to realize how much carbon being fixed in our system. We have to understand the process in E. coli after uptaking CO2. Since we integrated non-native carbon fixation pathway into E. coli to let E. coli utilize CO2, we simplify the CO2 utilization pathway in engineered E. coli into two parts, CO2 uptake and CO2 metabolism.
Method of model
Follow our model progress diagram with a better sense of how we achieve our goal.
Purpose of model
How many CO2 can be used by E. coli
CO2 uptake
CO2 exists in the phase of gas. Therefore, E. coli uptake CO2 by diffusion. However, the native diffusion rate is really low, not to mention the dissolved rate. We clone CA (carbonic anhydrase) gene to speed up the diffusion rate. The higher diffusion rate, the higher CO2 concentration maintained in engineered E. coli. Higher CO2 concentration in engineered E. coli provides more steady CO2 condition for downstream pathway.
| CO2 uptake | Kf (mM) | Kr (mM) | |||||
|---|---|---|---|---|---|---|---|
| R1 | $${[CO_2]_{air} \rightarrow [CO_2]_{uptake}}$$ | 9.23E-04 | 4.78E-04 | ||||
| R2 | $${[CO_2]_{uptake} + [H_2O] \rightarrow [H_2CO_3]}$$ | 0.062 | 23.7 | ||||
| R3 | $${[H_2CO_3] \rightarrow [H^+] + [HCO_3^-]}$$ | 8000000 | 4.7E10 | ||||
| R4 | $${[HCO_3^-] \rightarrow [H^+] + [CO_3^{2-}]}$$ | 3 | 5E10 | ||||
| R5 | $${[H^+] + [HCO_3^-] \longleftrightarrow [H_2O] + [CO_2]_{uptake}}$$ | Vmax | Km | ||||
| 0.47333 | 0.0062 | ||||||
Result
Enzyme, CA, strongly catalyzes the hydration of CO2 and its interaction appears to follow a Michaelis-Menten mechanism with a Hill constant of 0.0062 in our research. The result of Engineered E. coli cloned with or without CA gene showed below. A quite different diffusion time explains the function of CA.
Fig 1. CO2 uptake amount change with time in engineered E. coli without CA gene.
Fig 2. CO2 uptake amount change with time in engineered E. coli with CA gene.
CO2 Metabolism
Xylose is the main source of our engineered E. coli to uptake CO2. However, there is another pathway for xylose metabolism, which means that there is only a part of xylose that E. coli consumed will react with CO2. We build a model to predict the different carbon flux between original xylose metabolism, PP pathway (Pentose Phosphate pathway), and our CO2 bypass pathway.
Both of two pathways will end up at pyruvate, which is a key intersection in the network of metabolic pathways. We built a model of TCA cycle to explain how pyruvate play an energy precursor role and then is converted into biomass of E. coli.
Pathway
Fig 3. Metabolic pathway of CO2-utilization E. coli.
The introduced CO2-utilization bypass pathway composed of PRK and Rubisco is drawn in green line and noted by ” A” reaction and the double line was the pathway that composed by genetically modified, while the central carbon metabolic pathway including PP pathway and TCA cycle is drawn in blue line and yellow line and noted by “B” reaction and “C” reaction, respectively. See more detail in Kinetic law.
| CO2 bypass pathway | Vmax (1/S) | KM (mM) | |||||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| A1 | $${Xylose \rightarrow Xylulose}$$ $${Xylulose + ATP \rightarrow X5P}$$ | 0.02456 | 124.55879 | ||||||||||
| A2 | $${X5P \rightarrow Ru5P}$$ | 0.08 | 12.5 | ||||||||||
| A3 | $${Ru5P \xrightarrow{PRK} RuBP}$$ | 225.81858 | 0.27 | ||||||||||
| A4 | $${RuBP + CO_2 \xrightarrow{RuBisCO} 3PG}$$ | Vmax | Krubp | KCO2 | |||||||||
| 333.33 | 53 | 2.7 | |||||||||||
| Pentose Phosphate Pathway | Vmax (1/S) | KM (mM) | ||||||||||||||||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| B1 | $${Ru5P \rightarrow R5P}$$ | 0.13672 | 12.5 | |||||||||||||||||||||
| B2 | $${X5P + R5P \rightarrow S7P + GAP}$$ | |||||||||||||||||||||||
| $${X5P \rightarrow S7P}$$ | 58.27 | 4.96E-09 | ||||||||||||||||||||||
| $${X5P \rightarrow GAP}$$ | 58.27 | 48 | ||||||||||||||||||||||
| $${R5P \rightarrow S7P}$$ | 58.27 | 0.008 | ||||||||||||||||||||||
| $${R5P \rightarrow GAP}$$ | 58.27 | 3.05E-07 | ||||||||||||||||||||||
| B3 | $${S7P + GAP \rightarrow F6P + E4P}$$ | |||||||||||||||||||||||
| $${S7P \rightarrow E4P}$$ | 58.27 | 2.524 | ||||||||||||||||||||||
| $${S7P \rightarrow F6P}$$ | 58.27 | 6.10E-09 | ||||||||||||||||||||||
| $${GAP \rightarrow E4P}$$ | 58.27 | 1.73E-07 | ||||||||||||||||||||||
| $${GAP \rightarrow F6P}$$ | 58.27 | 8.23E-09 | ||||||||||||||||||||||
| B4 | $${E4P + X5P \rightarrow F6P + GAP}$$ | |||||||||||||||||||||||
| $${E4P \rightarrow F6P}$$ | 1 | 1 | ||||||||||||||||||||||
| $${E4P \rightarrow GAP}$$ | 5.827E-4 | 1.733E-7 | ||||||||||||||||||||||
| $${X5P \rightarrow F6P}$$ | 0.05827 | 1.733E-7 | ||||||||||||||||||||||
| $${X5P \rightarrow GAP}$$ | 0.005827 | 48.8 | ||||||||||||||||||||||
| B5 | $${FBP \rightarrow GAP}$$ | 135.425 | 4 | |||||||||||||||||||||
| B6 | $${DHAP \longleftrightarrow GAP}$$ | Vmf | Vmr | KmDHAP | KmGAP | |||||||||||||||||||
| 0.2394 | 0.24 | 0.812 | 0.245 | |||||||||||||||||||||
| B7 | $${FBP \rightarrow DHAP}$$ | 0.0001247 | 0.04 | |||||||||||||||||||||
| B8 | $${F6P + ATP \rightarrow FBP + ADP}$$ | 88.2533 | 0.017 | |||||||||||||||||||||
| B9 | $${F6P \rightarrow G6P}$$ | 88.2353 | 0.017 | |||||||||||||||||||||
| B10 | $${G6P + NADP^+ \rightarrow 6PG + NADPH}$$ | Vmax | KmG6P | KmNADP | ||||||||||||||||||||
| 11.736 | 0.12 | 0.0123 | ||||||||||||||||||||||
| B11 | $${6PG + NADP^+ \rightarrow Ru5P + NADPH}$$ | 1.0 | 0.044 | |||||||||||||||||||||
| B12 | $${GAP + NADP^+ + ADP \rightarrow 3PGA + NADPH + ATP}$$ | 0.4 | 0.013 | |||||||||||||||||||||
| B13 | $${3PGA \rightarrow PEP}$$ | 0.43478 | 0.23 | |||||||||||||||||||||
| B14 | $${PEP \rightarrow PYR}$$ | 0.23148 | 0.45 | |||||||||||||||||||||
| TCA cycle | Vmf (mM/min) | Vmr (mM/min) | Ks (mM) | Kp (mM) | ||||||||||||||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| C1 | $${PEP + HCO_3^- + NAD^+ \rightarrow OAA + NADH}$$ | 1 | 1 | 1 | 1 | |||||||||||||||||||
| C2 | $${AcCoA \rightarrow CIT}$$ | 64.8 | 0.648 | 0.05 | 0.12 | |||||||||||||||||||
| C3 | $${CIT \rightarrow ICIT}$$ | 31.2 | 0.312 | 1.7 | 0.7 | |||||||||||||||||||
| C4 | $${ICIT + NADP^+ \rightarrow AKG + NADPH + CO_2}$$ | 10.2 | 0.102 | 0.03 | 0.3 | |||||||||||||||||||
| C5 | $${AKG + NAD^+ \rightarrow AKG + NADPH + CO_2}$$ | 57.344 | 0.57344 | 0.1 | 1.0 | |||||||||||||||||||
| C6 | $${SUCCoA + ADP \rightarrow SUCC + ATP}$$ | 6.54 | 0.0651 | 0.015 | 0.15 | |||||||||||||||||||
| C7 | $${SUCC + FAD \rightarrow FUM + FADH_2}$$ | 1.02 | 1.02 | 0.12 | 0.15 | |||||||||||||||||||
| C8 | $${FUM \rightarrow MAL}$$ | 887.7 | 87.7 | 0.25 | 2.38 | |||||||||||||||||||
| C9 | $${MAL + NAD^+ \rightarrow OAA + NADH}$$ | 184 | 184 | 0.833 | 0.0443 | |||||||||||||||||||
| C10 | $${OAA \rightarrow CIT}$$ | 64.8 | 0.648 | 0.012 | 0.12 | |||||||||||||||||||
| C11 | $${PYR \rightarrow NADH + AcCoA + CO_2}$$ | Vmax (1/s) | Km (mM) | |||||||||||||||||||||
| 0.00518 | 1.5 | |||||||||||||||||||||||
| C12 | $${ICIT \rightarrow GOX}$$ | 1.172 | 0.01172 | 0.145 | 0.13 | |||||||||||||||||||
| C13 | $${GOX \rightarrow MAL}$$ | 20 | 0.2 | 0.057 | 1.0 | |||||||||||||||||||
| C14 | $${ICIT \rightarrow SUCC + GOX}$$ | Vmf (mM/min) | Vmr (mM/min) | Kicit (mM) | Ksucc (mM) | Kgox (mM) | ||||||||||||||||||
| 1.172 | 0.012 | 0.145 | 0.59 | 0.13 | ||||||||||||||||||||
| E4P | erythrose-4-phosphate |
| F6P | fructose-6-phosphate |
| FBP | fructose-1,6-bisphosphate |
| G6P | glucose-6-phosphate |
| GAP | glyceraldehyde-3-phosphate |
| PEP | phosphoenol pyruvate |
| PYR | pyruvate |
| R5P | ribose-5-phosphate |
| Ru5P | ribulose-5-phosphate |
| S7P | sedoheptulose-7-phosphate |
| X5P | xylulose-5-phosphate |
| 6PG | 6-phosphogluconate |
| DHAP | Dihydroxyacetone phosphate |
| 3PG | 3-Phosphoglyceric acid |
| RuBP | ribulose-1,5-bisphosphate |
| PRK | Phosphoribulokinase |
| Rubisco | Ribulose-1,5-bisphosphate carboxylase/ oxygenase |
| AcCoA | Acetyl-CoA |
| CIT | Citrate |
| ICIT | Isocitrate |
| AKG | Alpha-ketoglutarate |
| SUCCoA | Succinyl-CoA |
| SUCC | Succinate |
| FUM | Fumarate |
| MAL | Malate |
| OAA | Oxaloacetic acid |
| GOX | Glyoxylate |
Xylose v.s. CO2 analysis in CO2 utilization efficiency
Two main sources of CO2 metabolism in engineered E. coli are xylose and CO2. CO2 utilization rate varies under different condition. We use 5% CO2 (about 2.6 mM) and 4 (g/l) xylose (about 26 mM) in experiment as an optimization CO2 utilization rate. Constant CO2 condition and limited CO2 condition also effects metabolism performance. In other words, open system and close system showed different results. Therefore, adjust initial concentration of CO2 and Xylose and choose open or close system to see the change of pyruvate production.
References
- E. G. Jacqueline, P. L. Christopher, R. A. Maciek, Comprehensive analysis of glucose and xylose metabolism in Escherichia coli under aerobic and anaerobic conditions by 13C metabolic flux analysis. Metab Eng. 2017 Jan; 39: 9–18.
- U. Sauer, J. E. Bernhard, The PEP—pyruvate—oxaloacetate node as the switch point for carbon flux distribution in bacteria. FEMS Microbiology Reviews, Volume 29, Issue 4, 1 September 2005, Pages 765–794.
- G. Fuyu, L. Guoxia, Z. Xiaoyun, Z. Jie, C. Zhen, L. Yin . Quantitative analysis of an engineered CO2-fixing Escherichia colireveals great potential of heterotrophic CO2 fixation. Gong et al. Biotechnology for Biofuels, 2015, 8:86.
- Y. Pocker, S. Y. Ng. Joan, Plant carbonic anhydrase. Properties and carbon dioxide hydration kinetics. Biochemistry, 1973, 12 (25), pp 5127–5134.