Team:Evry/Project FUR

From 2013.igem.org

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<h1>FUR system</h1>
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<h1>Iron chelators</h1>
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<h2>FUR system</h2>
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<img src="https://static.igem.org/mediawiki/2013/b/ba/FUR-protein.png" width=200px/>
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<p>
<p>
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Iron is an essential element in the development of <i>E. coli</i>, but also, it can be toxic and <i>E. coli</i> can be killed, if iron is absorbed in high quantity. Using the ferric-uptake regulator protein (Fur), bacteria developed an advanced system to regulate their iron homeostasis.
+
Iron is an essential element in the development of <i>E. coli</i>, but it can also be toxic and <i>E. coli</i> can be killed, if iron is absorbed in high quantity. Using the ferric-uptake regulator protein (Fur), bacteria developed an advanced system to regulate their iron homeostasis.
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<p>
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The Fur protein is a transcriptional repressor for more than 90 genes involved, in majority, in iron homeostasis<sup>1,2</sup>. It plays an important role in the control of the intracellular concentration of iron in <i>E. coli</i>.
+
The Fur protein acts a <b>transcriptional repressor</b> for more than 90 genes involved, in majority, in <b>iron homeostasis</b><sup>1,2</sup>. It plays an important role in the control of the intracellular concentration of iron in <i>E. coli</i>.
</p>
</p>
<p>
<p>
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Fur acts as a positive repressor in presence of ferrous ion (Fe<sup>2+</sup>), its co-repressor. Then, Fe<sup>2+</sup> binds to the Fur protein (one ferrous ion per subunit of Fur). This interaction leads to a structural modification and induces the dimerization of Fur and Fe<sup>2+</sup>. Then the homodimeric Fur-Fe<sup>2+</sup> complex will bind to the DNA on a Fur binding site and inhibit the mRNA transcription. In absence of Fe<sup>2+</sup>, a disinhibiting effect occurs and mRNA can be transcribed.
+
Fur acts as an <b>active repressor</b> in presence of ferrous ion (Fe<sup>2+</sup>), its co-repressor. Then, Fe<sup>2+</sup> binds to the Fur protein (one ferrous ion per subunit of Fur). This interaction leads to a structural modification and induces the dimerization of Fur and Fe<sup>2+</sup>. Then the homodimeric Fur-Fe<sup>2+</sup> complex binds to the DNA on a Fur binding site and inhibits the mRNA transcription. In absence of Fe<sup>2+</sup>, a disinhibiting effect occurs and mRNA can be transcribed.
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   <b>Figure 1:</b> Regulation by the Fur transcriptional factor. In absence of iron mRNA can be transcribed. In presence of iron, Fur binds the DNA, on the Fur binding site and mRNA expression in inhibited.
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   <b>Figure 1:</b> Regulation by the Fur transcriptional factor. In absence of iron mRNA can be transcribed. In presence of iron, Fur binds the DNA, on the Fur binding site and mRNA expression is inhibited.
  </div>
  </div>
</div>
</div>
<p>
<p>
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Fur is found at a level of 5000 copies in E. coli cell during exponential growth and it can be up to 10’000 copies in stationary phase. Note that this high number of Fur protein is essential to E. coli during its development, because regulating the expression of gene involved in the iron absorption, it avoids an over absorption of iron which could be really toxic and kill E. coli.  
+
Fur is found at a level of 5000 copies in <i>E. coli</i> during exponential growth and it can reach up to 10,000 copies in stationary phase. Note that this high number of Fur protein is essential to <i>E. coli</i> during its development. As explained previously, Fur regulates the expression of genes involved in iron absorption, and it avoids an over absorption of iron which could be really toxic and kill <i>E. coli</i>.  
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   <b>Figure 2:</b> Légende ici.
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   <b>Figure 2:</b> Consensus palindromic sequence of Fur binding site.
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<p style="padding-top:1%;">
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The <b>Fur Binding</b> site also named <b>“Fur box”</b> or <b>“iron box”</b> is localized between -35 and -10 site at the promoters of Fur-repressed genes. As shown in the Figure 2, the Fur binding site is composed of 19 bases pair that are organized as a <b>palindromic consensus sequence</b>. It must be noted that this consensus sequence is not an exact sequence. Others sequences of Fur binding sites can be found into E. coli’s genome.
+
The <b>Fur Binding</b> site also named <b>“Fur box”</b> or <b>“iron box”</b> is localized between -35 and -10 site at the promoters of Fur-repressed genes. As shown in the Figure 2, the Fur binding site is composed of 19-base pair that are organized as a <b>palindromic consensus sequence</b>. It must be noted that this consensus sequence is not an exact sequence. Others sequences of Fur binding sites can be found into <i>E. coli</i>’s genome.
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<p>
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Often, Fur binding site is repeated (as shown in the Figures 3 and 4). It means that several homodimer of Fur-Fe<sup>2+</sup> can bind among the DNA. Then Fur binding sites can be localized in a larger region than between -35 and -10 of the promoter region (defined previously). The affinity of these Fur binding site to Fur-Fe<sup>2+</sup> is different. In a first time the complex Fur-Fe<sup>2+</sup> binds to the 1<sup>st</sup> Fur binding site, which presents a higher affinity for its complex. Then, others Fur-Fe<sup>2+</sup> bind to the following Fur binding sites which present a weaker affinity. A polymer of Fur protein is formed among the DNA which allows a stronger inhibition of the expression of the gene downstream these Fur binding sites.
+
Often, Fur binding site is repeated (as shown in the Figures 3 and 4). It means that several homodimer of Fur-Fe<sup>2+</sup> can bind among the DNA. Then Fur binding sites can be localized in a larger region than between -35 and -10 of the promoter region (defined previously). The affinity of these Fur binding site to Fur-Fe<sup>2+</sup> is different. At first the complex Fur-Fe<sup>2+</sup> binds to the 1<sup>st</sup> Fur binding site, which presents a higher affinity for its complex. Then, others Fur-Fe<sup>2+</sup> bind to the following Fur binding sites which present a weaker affinity. A polymer of Fur protein is formed among the DNA which allows a stronger inhibition of the expression of the gene downstream these Fur binding sites.
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   <b>Figure 3:</b> Légende ici.
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   <b>Figure 3:</b> Repeated Fur binding site sequences.
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   <b>Figure 4:</b> Légende ici.
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   <b>Figure 4:</b> Repeated Fur binding site sequences on a double strand DNA.
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<p>
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It had been shown that Fur could be involved in a positive regulation of several genes of E. coli (<i>fntA</i>, <i>bfr</i>, <i>acnA</i>, <i>fumA</i>, <i>sdh</i> operon, <i>sodB</i>). In fact E. coli uses an inverter system mediated by a small RNA named RhyB.<br/>
+
It had been shown that Fur could be involved in a positive regulation of several genes of <i>E. coli</i> (<i>fntA</i>, <i>bfr</i>, <i>acnA</i>, <i>fumA</i>, <i>sdh</i> operon, <i>sodB</i>). In fact <i>E. coli</i> uses an inverter system mediated by a small RNA named RhyB.<br/>
RhyB is a 90 nucleotids long sRNA which is regulated by the Fur transcriptional factor, then it is expressed at low concentration of iron<sup>3</sup>.
RhyB is a 90 nucleotids long sRNA which is regulated by the Fur transcriptional factor, then it is expressed at low concentration of iron<sup>3</sup>.
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   <b>Figure 5:</b> Légende ici.
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   <b>Figure 5:</b> Regulation by the Fur transcriptional and the RhyB sRNA, acting as an inverter. In absence of iron mRNA is binded by the RhyB and then it is degradated. In presence of iron, Fur binds the DNA, the RhyB expression is inhibited and the mRNA can be transcribed.
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<p>
As shown in Figure 5, and if we apply it to the sodB gene expression system. <b>In iron starvation</b>, Fur-Fe<sup>2+</sup> cannot be formed and RyhB transcription is disinhibited. RyhB is expressed in the intracellular environment and it can binds to sodB mRNA which contains RyhB’s target sequence. Once RyhB binds to SodB mRNA, RNA degrading enzymes, such as RNaseE and RNase III, are recruited and <b>the new formed complex is digested.</b><br/>
As shown in Figure 5, and if we apply it to the sodB gene expression system. <b>In iron starvation</b>, Fur-Fe<sup>2+</sup> cannot be formed and RyhB transcription is disinhibited. RyhB is expressed in the intracellular environment and it can binds to sodB mRNA which contains RyhB’s target sequence. Once RyhB binds to SodB mRNA, RNA degrading enzymes, such as RNaseE and RNase III, are recruited and <b>the new formed complex is digested.</b><br/>
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However, if the iron concentration high enough Fur-Fe<sup>2+</sup> complex is formed and it can inhibit the RyhB transcription. SodB is not repressed and can be synthesized<sup>3</sup>. That is why Fur is described as an activator transcriptional factor in such a case.
+
However, if the iron concentration is high enough, Fur-Fe<sup>2+</sup> complex is formed and it can inhibit the RyhB transcription. SodB is not repressed and can be synthesized<sup>3</sup>. That is why Fur is described as an activator transcriptional factor in such a case.
</p>
</p>
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<h3>Siderophores</h3>
<h3>Siderophores</h3>
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<p>Siderophores are molecules produces by bacteria to get iron when there is a low amount of it in the environment. There is different types of siderophores, depending of the nature of their bounds with iron: Catecholate, phenolate, hydroxymate or carboxylate types.<br/>
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<p>
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Enterobactins are catecholate siderophore (composed with catechol = bi-phenol) with very high affinity for ferric iron K = 10<sup>52</sup> M<sup>-1</sup>. This number is greater than the affinity of b heme (compound of human hemo and myoglobins) for iron (K= 10<sup>39</sup> M<sup>−1</sup>) or  the affinity of EDTA metal chelatorfor iron (10<sup>25</sup> M<sup>−1</sup>). De facto, enterobactin is one of the powerfull siderophore known: it can extract iron from hemic source and even from air !</p>
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During iron starvation, bacteria produce siderophores, small molecules that can catch any atom of iron in the environment to feed the bacteria. There are different types of siderophores, depending on their structure and their interaction with iron:
 +
</p>
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<ul>
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<li>Catecholate,</li>
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<li>phenolate,</li>
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<li>hydroxymate,</li>
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<li>carboxylate types.</li>
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</ul>
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<p>
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We focus more specifically on <b>enterobactin</b> which is a catecholate siderophore (composed of catechol = bi-phenol) produced by <i>E. coli</i>. Enterobactin has a very high affinity for ferric iron K = 10<sup>52</sup> M<sup>-1</sup><sup>4</sup>. This number is greater than the affinity of heme B and EDTA metal chelator for iron, respectively K = 10<sup>39</sup> M<sup>−1</sup> and 10<sup>25</sup> M<sup>−1</sup>. <i>De facto</i>, enterobactin is one of the most powerfull siderophores known: it can extract iron from hemic source and even from air!
 +
</p>
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   <b>Figure 7:</b> Structure of Enterobactin molecule  
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   <b>Figure 7:</b> Structure of Enterobactin molecule.
  </div>
  </div>
</div>
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</div>
<br/>
<br/>
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<p>Ferric iron Fe<sup>3+</sup> established hydrogen bound with the alcohol groupment of enterobactins.</p>
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<p>Ferric iron Fe<sup>3+</sup> or Ferrous iron Fe<sup>2+</sup> established hydrogen bound with the alcohol groupment of enterobactin.</p>
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   <b>Figure 8:</b> Hydrogen bounds with Ferric Iron </br> (from Raymond, K. N. 2003; <a href="http://www.pnas.org/content/100/7/3584.full.pdf" target=‘_blank'>link to the pdf</a>).
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   <b>Figure 8:</b> 3D structure of enterobaction without (A) or with (B) Ferric Iron<sup>5</sup>.
  </div>
  </div>
</div>
</div>
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<p>Enterobactin pathway is under the regulation of the Fur System in <i>E.coli</i>. When there is a lack of iron in the environment, genes of enterobactin pathway are expressed.<br/>
 
-
In our project, we focus on the 6 enzymes (EntA, EntB, EntC, EntD, EntE and EntF) of it biosynthesis from chorismic acid but there are other genes involved in this pathway as FepA that export enterobactin out of the bacteria.<br/>
 
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The biosynthesis pathway of enterobactin is the following:
 
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<div align="center"><img src="https://static.igem.org/mediawiki/2013/3/3b/EntBS.png"/ width="100%"></div>
 
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<p><i><small>Note: The last step  of the biosynthesis remains unclear.</small></i></p>
 
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<h3>Constructions</h3>
 
<p>
<p>
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Our goal is to lower the iron absorption from the intestins to the blood by using an iron chelating bacteria, <b>Iron Coli</b>. The objective of the labwork is to design an iron-sensible promoter with FUR and overexpress the enterobactin synthesis pathway in the presence of iron.
+
Genes of enterobactin pathway are under the regulation of the Fur protein in <i>E.coli</i>. It means that genes of enterobactin pathway are expressed only in lack of iron.
</p>
</p>
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<div align="center">
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<div class="captionedPicture" style="width:90%;float:center;">
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<a title="Enterobactines Synth" href="https://static.igem.org/mediawiki/2013/3/3b/EntBS.png">
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  <img alt="Enterobactine Synths" src="https://static.igem.org/mediawiki/2013/3/3b/EntBS.png" class="Picture"/>
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</a>
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<div class="caption">
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  <b>Figure 9:</b> Pathway of the synthese of the enterobactin.
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</div>
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</div>
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</div>
<p>
<p>
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Before going in the detail of our construction, we needed a iron marker in <i>E. coli</i>. Thus, we selected the FUR protein which negatively regulates the downstream genes. It is used in the iron regulation of the bacterium and smoothly shuts down the production ot siderophores, natural chelators of iron in the environment in iron depletion situation. When the concentration of iron exceeds 10<sup>6</sup>, the FUR proteins binds the ion, dimerizes and its inhibitory mechanism is activated. Below this threshold, the FUR protein's inhibition is suppressed and indirectly activates the downstream genes.
+
In our project, we want to produce enterobactin. Then we focus on the 6 enzymes (EntA, EntB, EntC, EntD, EntE and EntF) of it biosynthesis started from chorismic acid. However, we want to produce enterobactin in presence of iron, in order to reduce an excess of iron.<br/>
</p>
</p>
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 +
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<h2 id="FbpA">An alternative iron chelator</h2>
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<p>At the Regional Jamboree, we discussed with <a ref="https://2013.igem.org/Team:Edinburgh" target=‘_blank'>Edinburgh team</a> which also work on iron chelation but with another chelator: the ferric ion-binding protein A (FbpA)<sup><small>6</sup></small>. <br/>
 +
This protein from <i>Neisseria gonorrhoeae</i> is able to chelate various ion in solution as Fe<sup><small>3+</sup></small>
 +
, Ti<sup><small>4+</sup></small>,Cu<sup><small>2+</sup></small>, Zr<sup><small>4+</sup></small>(Zirconium ion)
 +
or Hf<sup><small>4+</sup></small> (Hafnium ion)<sup><small>7</sup></small>.</br>
 +
Its affinity for ferrous iron is 10<sup><small>18</sup></small>, less than our enterobactins system, but it remains a good alternative for our treatment.</p>
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 +
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<h2>Constructions</h2>
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   <b>Figure 6:</b> Construction..
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   <b>Figure 10:</b> Enterobactine production using the Fur-inverter system.
  </div>
  </div>
</div>
</div>
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<h3>Conclusion</h3>
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<p>
-
 
+
Our goal is to decrease the iron absorption from the intestines to the blood by using an iron chelating bacteria, <b>Iron Coli</b>.<br/>
-
<p>In our project, we have design a system that could produce <a href="https://2013.igem.org/Team:Evry/Chelator" target=‘_blank'>enterobactins</a> from chorimsic acid, under the regulation of an iron <a href="https://2013.igem.org/Team:Evry/Sensor" target=‘_blank'>iron sensor</a> with an <a href="https://2013.igem.org/Team:Evry/Inverter" target=‘_blank'>inverter</a>. Those, when the amount in the environment (<i>ie</i> intestine) is high, our <b><span style="color:#bb8900">Iron</span><span style="color:#7B0000"> Coli</span></b> will produce lot of siderophores that will chelate the iron and hence reduce the amount that could be absorbed by the patient.<br/>
+
We have design a system that could produce <a href="https://2013.igem.org/Team:Evry/Chelator" target=‘_blank'>enterobactin</a> from chorimsic acid, under the regulation of an <a href="https://2013.igem.org/Team:Evry/Sensor" target=‘_blank'>iron sensor</a> with an <a href="https://2013.igem.org/Team:Evry/Inverter" target=‘_blank'>inverter</a>. When the iron is present in the environment (<i>ie</i> intestine) in high concentration, our <b><span style="color:#bb8900">Iron</span><span style="color:#7B0000"> Coli</span></b> will produce siderophores to chelate the iron and hence reduce the amount of iron that could be absorbed by the patient.<br/>
-
We also model <a href="https://2013.igem.org/Team:Evry/Metabolism_model" target=‘_blank'>enterobactin production</a> of our <b><span style="color:#bb8900">Iron</span><span style="color:#7B0000"> Coli</span></b> and the potential <a href="https://2013.igem.org/Team:Evry/Model1" target=‘_blank'>efficiency of our treatment</a> to cure iron overload.</p>  
+
We also model <a href="https://2013.igem.org/Team:Evry/Metabolism_model" target=‘_blank'>enterobactin production</a> of our <b><span style="color:#bb8900">Iron</span><span style="color:#7B0000"> Coli</span></b> and the potential <a href="https://2013.igem.org/Team:Evry/Model1" target=‘_blank'>efficiency of our treatment</a> to cure iron overload.
-
 
+
</p>
<div id="citation_box">
<div id="citation_box">
-
  <p id="references">References:</p>
+
  <p id="references"><b>References:</b></p>
  <ol>
  <ol>
-
   <li>Revue</li>
+
   <li>Andrews, S. C., Robinson, A. K. & Rodriguez-Quiñones, F. Bacterial iron homeostasis. <i>FEMS Microbiology Reviews</i> 27, 215–237 (2003).</li>
-
   <li>“Iron and metal regulation in bacteria”, Klaus Hantke)</li>
+
   <li>Klaus Hantke, Iron and metal regulation in bacteria, <i>Elsevier</i>,2001, vol. 4, no2, pp. 172-177</li>
-
   <li>(ref: Amanda G., “Iron responsive Bacterial small RNAs: variation on a theme”)</li>
+
   <li>Amanda G., Iron responsive Bacterial small RNAs: variation on a theme, <i>Metallomics</i>, January 2013</li>
-
</ol>
+
  <li>Zheng, T., Siderophore-Mediated Cargo Delivery to the Cytoplasm of Escherichia coli and Pseudomonas aeruginosa : Syntheses of Monofunctionalized Enterobactin Scaffolds and Evaluation of Enterobactin–Cargo Conjugate Uptake, <i>J Am Chem Soc.</i>, 2012 Nov 7.
 +
  <li> Raymond, K. N., Enterobactin: An archetype for microbial iron transport, <i>PNAS</i>, vol.100, n°7, April 2003.</li>
 +
  <li>Ferreiros, C., Criado, M. and Gomez, J. (1999) The Neisserial 37kDa ferric binding protein (FbpA). Comparative Biochemistry and Physiology Part B. 123. 1-7. </li>
 +
  <li>Alexeev, D., Zhu, H., Guo, M., Zhong, W., Hunter, D., Yang, W., Campopiano, D. and Sadler, P. (2003) A novel protein-mineral interface. Nature Structural Biology. 10. 297-302. </li>
 +
</ol>
 +
 
</div>
</div>

Latest revision as of 03:23, 29 October 2013

Iron coli project

Iron chelators

FUR system



Iron is an essential element in the development of E. coli, but it can also be toxic and E. coli can be killed, if iron is absorbed in high quantity. Using the ferric-uptake regulator protein (Fur), bacteria developed an advanced system to regulate their iron homeostasis.

FUR protein (Ferric Uptake Regulator)

The Fur protein acts a transcriptional repressor for more than 90 genes involved, in majority, in iron homeostasis1,2. It plays an important role in the control of the intracellular concentration of iron in E. coli.

Fur acts as an active repressor in presence of ferrous ion (Fe2+), its co-repressor. Then, Fe2+ binds to the Fur protein (one ferrous ion per subunit of Fur). This interaction leads to a structural modification and induces the dimerization of Fur and Fe2+. Then the homodimeric Fur-Fe2+ complex binds to the DNA on a Fur binding site and inhibits the mRNA transcription. In absence of Fe2+, a disinhibiting effect occurs and mRNA can be transcribed.

Meca_FurBS
Figure 1: Regulation by the Fur transcriptional factor. In absence of iron mRNA can be transcribed. In presence of iron, Fur binds the DNA, on the Fur binding site and mRNA expression is inhibited.

Fur is found at a level of 5000 copies in E. coli during exponential growth and it can reach up to 10,000 copies in stationary phase. Note that this high number of Fur protein is essential to E. coli during its development. As explained previously, Fur regulates the expression of genes involved in iron absorption, and it avoids an over absorption of iron which could be really toxic and kill E. coli.

FUR binding site architecture

Meca_RyhB
Figure 2: Consensus palindromic sequence of Fur binding site.

The Fur Binding site also named “Fur box” or “iron box” is localized between -35 and -10 site at the promoters of Fur-repressed genes. As shown in the Figure 2, the Fur binding site is composed of 19-base pair that are organized as a palindromic consensus sequence. It must be noted that this consensus sequence is not an exact sequence. Others sequences of Fur binding sites can be found into E. coli’s genome.

Often, Fur binding site is repeated (as shown in the Figures 3 and 4). It means that several homodimer of Fur-Fe2+ can bind among the DNA. Then Fur binding sites can be localized in a larger region than between -35 and -10 of the promoter region (defined previously). The affinity of these Fur binding site to Fur-Fe2+ is different. At first the complex Fur-Fe2+ binds to the 1st Fur binding site, which presents a higher affinity for its complex. Then, others Fur-Fe2+ bind to the following Fur binding sites which present a weaker affinity. A polymer of Fur protein is formed among the DNA which allows a stronger inhibition of the expression of the gene downstream these Fur binding sites.

Meca_RyhB
Figure 3: Repeated Fur binding site sequences.
Meca_RyhB
Figure 4: Repeated Fur binding site sequences on a double strand DNA.

Natural inverter system

It had been shown that Fur could be involved in a positive regulation of several genes of E. coli (fntA, bfr, acnA, fumA, sdh operon, sodB). In fact E. coli uses an inverter system mediated by a small RNA named RhyB.
RhyB is a 90 nucleotids long sRNA which is regulated by the Fur transcriptional factor, then it is expressed at low concentration of iron3.

Meca_RyhB
Figure 5: Regulation by the Fur transcriptional and the RhyB sRNA, acting as an inverter. In absence of iron mRNA is binded by the RhyB and then it is degradated. In presence of iron, Fur binds the DNA, the RhyB expression is inhibited and the mRNA can be transcribed.

As shown in Figure 5, and if we apply it to the sodB gene expression system. In iron starvation, Fur-Fe2+ cannot be formed and RyhB transcription is disinhibited. RyhB is expressed in the intracellular environment and it can binds to sodB mRNA which contains RyhB’s target sequence. Once RyhB binds to SodB mRNA, RNA degrading enzymes, such as RNaseE and RNase III, are recruited and the new formed complex is digested.
However, if the iron concentration is high enough, Fur-Fe2+ complex is formed and it can inhibit the RyhB transcription. SodB is not repressed and can be synthesized3. That is why Fur is described as an activator transcriptional factor in such a case.

Siderophores

During iron starvation, bacteria produce siderophores, small molecules that can catch any atom of iron in the environment to feed the bacteria. There are different types of siderophores, depending on their structure and their interaction with iron:

  • Catecholate,
  • phenolate,
  • hydroxymate,
  • carboxylate types.

We focus more specifically on enterobactin which is a catecholate siderophore (composed of catechol = bi-phenol) produced by E. coli. Enterobactin has a very high affinity for ferric iron K = 1052 M-14. This number is greater than the affinity of heme B and EDTA metal chelator for iron, respectively K = 1039 M−1 and 1025 M−1. De facto, enterobactin is one of the most powerfull siderophores known: it can extract iron from hemic source and even from air!

Enterobactines Structure
Figure 7: Structure of Enterobactin molecule.

Ferric iron Fe3+ or Ferrous iron Fe2+ established hydrogen bound with the alcohol groupment of enterobactin.

Enterobactines
Figure 8: 3D structure of enterobaction without (A) or with (B) Ferric Iron5.

Genes of enterobactin pathway are under the regulation of the Fur protein in E.coli. It means that genes of enterobactin pathway are expressed only in lack of iron.

Enterobactine Synths
Figure 9: Pathway of the synthese of the enterobactin.

In our project, we want to produce enterobactin. Then we focus on the 6 enzymes (EntA, EntB, EntC, EntD, EntE and EntF) of it biosynthesis started from chorismic acid. However, we want to produce enterobactin in presence of iron, in order to reduce an excess of iron.

An alternative iron chelator

At the Regional Jamboree, we discussed with Edinburgh team which also work on iron chelation but with another chelator: the ferric ion-binding protein A (FbpA)6.
This protein from Neisseria gonorrhoeae is able to chelate various ion in solution as Fe3+ , Ti4+,Cu2+, Zr4+(Zirconium ion) or Hf4+ (Hafnium ion)7.
Its affinity for ferrous iron is 1018, less than our enterobactins system, but it remains a good alternative for our treatment.

Constructions

Meca_RyhB
Figure 10: Enterobactine production using the Fur-inverter system.

Our goal is to decrease the iron absorption from the intestines to the blood by using an iron chelating bacteria, Iron Coli.
We have design a system that could produce enterobactin from chorimsic acid, under the regulation of an iron sensor with an inverter. When the iron is present in the environment (ie intestine) in high concentration, our Iron Coli will produce siderophores to chelate the iron and hence reduce the amount of iron that could be absorbed by the patient.
We also model enterobactin production of our Iron Coli and the potential efficiency of our treatment to cure iron overload.

References:

  1. Andrews, S. C., Robinson, A. K. & Rodriguez-Quiñones, F. Bacterial iron homeostasis. FEMS Microbiology Reviews 27, 215–237 (2003).
  2. Klaus Hantke, Iron and metal regulation in bacteria, Elsevier,2001, vol. 4, no2, pp. 172-177
  3. Amanda G., Iron responsive Bacterial small RNAs: variation on a theme, Metallomics, January 2013
  4. Zheng, T., Siderophore-Mediated Cargo Delivery to the Cytoplasm of Escherichia coli and Pseudomonas aeruginosa : Syntheses of Monofunctionalized Enterobactin Scaffolds and Evaluation of Enterobactin–Cargo Conjugate Uptake, J Am Chem Soc., 2012 Nov 7.
  5. Raymond, K. N., Enterobactin: An archetype for microbial iron transport, PNAS, vol.100, n°7, April 2003.
  6. Ferreiros, C., Criado, M. and Gomez, J. (1999) The Neisserial 37kDa ferric binding protein (FbpA). Comparative Biochemistry and Physiology Part B. 123. 1-7.
  7. Alexeev, D., Zhu, H., Guo, M., Zhong, W., Hunter, D., Yang, W., Campopiano, D. and Sadler, P. (2003) A novel protein-mineral interface. Nature Structural Biology. 10. 297-302.