Team:Bielefeld-CeBiTec/Results/CO2-fixation

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The particular aim of the second module is to implement the carbon dioxide fiaxtion in <i>E.&nbsp;coli</i>. For this approach all items, like the <a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Results/CO2-fixation/Calvin-Cycle" target="_blank">sedoheptulose-1,7-bisphosphatase</a>, the <a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Results/CO2-fixation/RuBisCO" target="_blank">ribulose-1,5-bisphosphate carboxylase/oxygenase</a>  (RuBisCO) and the mechanism of <a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Results/CO2-fixation/RuBisCO" target="_blank">carbon dioxide fixation</a> were tested separetly in various approaches. For the optimization of the carbon dioxide fixation under aerobic growth conditions we investigate the anerobic microcompartment from <i>Halothiobacillus&nbsp;neapolitanus</i>, called the <a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Results/CO2-fixation/Carboxysome" target="_blank">carboxysom</a>.
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The particular aim of the second module is to implement the carbon dioxide fiaxtion in <i>E.&nbsp;coli</i>. Therefore we selected the Calvin cycle (figure 1) and used a bottom up approach. All heterologous expressed components, like the <a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Results/CO2-fixation/Calvin-Cycle" target="_blank">sedoheptulose-1,7-bisphosphatase (<i>glpX</i>)</a>, the phosphoribulokinase (<i>prkA</i>) , the <a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Results/CO2-fixation/RuBisCO" target="_blank">ribulose-1,5-bisphosphate carboxylase/oxygenase</a>  (RuBisCO) were tested separately in various experiments. The RubisCO is known to function best under high CO<sub>2</sub> concentration. To accomplish optimal conditions for the RubisCO in a very local enviroment a microcompartiment from <i>Halothiobacillus&nbsp;neapolitanus</i>, which is called carboxysome, was constructed in <i>E.&nbsp;coli</i>.
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<font size="2" style="text-align:center;"><b>Figure 1:</b> Schematic representation of the Calvin cylce. The reaction shown in green can be catalyzed by enzymes that naturally exist in <i>E.&nbsp;coli</i>, while the red one need to be expressed heterologous to enable the whole <a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Results/CO2-fixation/Calvin-Cycle">Calvin cycle</a> in <i>E.&nbsp;coli</i>.</font>
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<font size="2" style="text-align:center;"><b>Figure 1:</b> Schematic representation of the Calvin cylce. The reactions shown in green can be catalyzed by enzymes that naturally exist in <i>E.&nbsp;coli</i>, while the red ones need to be expressed heterologous to enable the whole <a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Results/CO2-fixation/Calvin-Cycle">Calvin cycle</a> in <i>E.&nbsp;coli</i>.</font>
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For this purpose we made different approaches. First of all we identified two <a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Results/CO2-fixation/RuBisCO">RuBisCOs</a> (from <i>H.&nbsp;neapolitanus</i> and from <i>Synechococcus&nbsp;elongatus</i>) as the most efficient and best fitting ones. There coding sequences were synthesized to remove illegal restriction sites and to optimize the codong usage for the heterologous expression in <i>E.&nbsp;coli</i>. These enzymes are known to function best inside a carboxysome. Therefore we started the construction of such a microcompartment in <i>E.&nbsp;coli</i>.
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Latest revision as of 02:57, 18 October 2014


Module II - Carbon Dioxide (CO2) Fixation

The particular aim of the second module is to implement the carbon dioxide fiaxtion in E. coli. Therefore we selected the Calvin cycle (figure 1) and used a bottom up approach. All heterologous expressed components, like the sedoheptulose-1,7-bisphosphatase (glpX), the phosphoribulokinase (prkA) , the ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO) were tested separately in various experiments. The RubisCO is known to function best under high CO2 concentration. To accomplish optimal conditions for the RubisCO in a very local enviroment a microcompartiment from Halothiobacillus neapolitanus, which is called carboxysome, was constructed in E. coli.



Figure 1: Schematic representation of the Calvin cylce. The reactions shown in green can be catalyzed by enzymes that naturally exist in E. coli, while the red ones need to be expressed heterologous to enable the whole Calvin cycle in E. coli.

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