Team:Hong Kong CUHK/vs

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<div id="partable">
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<p2>
<p align="center"><img src="https://static.igem.org/mediawiki/2013/2/29/VS.JPG"/></p>
<p align="center"><img src="https://static.igem.org/mediawiki/2013/2/29/VS.JPG"/></p>
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<p2> <p>The voltage switch is a novel protein switch that responds to external voltage.  The switch itself consist of the PDZ Ligand-Voltage sensor peptide (<a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092007">BBa_K1092007</a>) and the PDZ  Domain-Voltage sensor peptide (<a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092008">BBa_K1092008</a>), and can be  linked to different effectors such as the Dioxygenase (<a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092002">BBa_K1092002</a>) and Laccase (<a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092004">BBa_K1092004</a>), or the RFP  fragments (<a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092105">BBa_K1092105</a> &amp; <a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092106">BBa_K1092106</a>). Initially,  the two proteins would express and localize onto the inner membrane of the  bacteria. The two peptides would then come together forming a dimer. After  that, due to the mutual repulsion of the positive charges in the two voltage  sensor peptide, the two voltage sensor peptide would separate (Fig. 1). This  separate the two effectors down below the two peptide, causing either a long  distance for the two effectors to interact, or a longer distance of diffusion  of substrates. This represent the OFF stage of the voltage switch, and it could  be enhanced by using negatively charged environment to further pull them apart.<br />
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<p><h4>Voltage Sensor Peptide (Protein Domain <a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092006">BBa_K1092006</a>)</h4></p>
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    <br />
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  <p align="center"><img src="https://static.igem.org/mediawiki/2013/6/6e/S1.png"></p>
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During the ON stage, the environment become positively charged, and this  environment exert a strong electrostatic repulsion on the two voltage sensor peptide  so that they can be pushed together by overcoming the mutual repulsion between  the peptides (Fig. 2). This action brings the two downstream effectors  together, which either provides short-enough distance for them to interact, or  greatly reduces the distance of diffusion of the substrates, thus greatly  enhance the reaction rate.</p>
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   <p align="left">
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  <h4>Voltage Switch (Protein Domain <a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092006">BBa_K1092006</a>)</h4>
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  Voltage sensor peptide, from <em>Aeropyrum pernix</em>, is a protein domain found in a transmembrane potassium ion channel. It contains positively-charged 4 arginine residues, and also numerous hydrophobic leucine residues. Study has shown that this helical voltage sensor peptide is capable of moving within the membrane under external voltage. To further adapt this voltage sensor peptide into our project, we added in glycine serine flexible linker, and polyproline rigid linker.</p>
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   <p align="left"><br />
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    It is a protein domain found in a transmembrane potassium ion channel from <em>Aeropyrum  pernix</em>, which is highly voltage dependent. It contain positive charges  from the 4 arginine residues, and also numerous leucine residues which give it  high hydrophobicity. All these together make it an excellent  voltage sensing peptide in the transmembrane region. Study has shown that this helical voltage sensor peptide is capable of moving across the membrane under external voltage, which is the key to the voltage dependence of the ion channel  (PMID: 12721618). We make use of this feature to construct a novel voltage switch (<a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092007">BBa_K1092007</a> &amp; <a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092008">BBa_K1092008</a>), which consist of the PDZ domain (<a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092102">BBa_K1092102</a>), PDZ ligand (<a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092101">BBa_K1092101</a>), this voltage sensor peptide itself. To further adapt this voltage sensor peptide into our project, we added in two (GGGGS)3 flexible linker which serve to link other protein domains, and a (PE)5 linker which serve as a rigid rod that project from the sensor peptide to elongate it  and increase the separation distance.</p>
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  <p>Part-only sequencing result: <br />
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    (                      (GGGGS)3  linker                         )(<br />
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    GGTGGTGGTG  GTTCTGGTGG TGGTGGTTCT GGTGGTGGTG GTTCTCTGGG <br />
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    Voltage sensor  peptide<br />
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    TCTGTTTCGT CTGGTGCGCC  TGCTGCGTTT TCTGCGTATT CTGCTGATTA <br />
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    )(                                      (Pe)5 linker           )(<br />
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    TTTGCCCGGA ACCGGAACCG  GAACCAGAAC CAGAAGGTGG TGGTGGTTCT <br />
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    (GGGGS)3 linker                           )<br />
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    GGTGGTGGTG GTTCTGGTGG  TGGTGGTTCT</p>
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   <p align="left">Note: part name should be voltage sensor peptide, not  voltage switch </p>
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   <p>&nbsp;</p>
   <p>&nbsp;</p>
   <h4>ssDsbA-PDZ Ligand-Voltage Switch (Protein Domain <a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092007">BBa_K1092007</a>)</h4>
   <h4>ssDsbA-PDZ Ligand-Voltage Switch (Protein Domain <a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092007">BBa_K1092007</a>)</h4>
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  <p align="center"><img src="https://static.igem.org/mediawiki/2013/2/24/Sh3.JPG"> </p>
   <p>It is a part of the voltage switch, which consist of the PDZ Ligand (<a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092101">BBa_K1092101</a>) and the voltage  sensor peptide (<a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092006">BBa_K1092006</a>). PDZ Ligand  can form dimers with PDZ domain (<a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092102">BBa_K1092102</a>), which links  this part together with the PDZ domain-Voltage sensor peptide (<a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092008">BBa_K1092008</a>).<br />
   <p>It is a part of the voltage switch, which consist of the PDZ Ligand (<a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092101">BBa_K1092101</a>) and the voltage  sensor peptide (<a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092006">BBa_K1092006</a>). PDZ Ligand  can form dimers with PDZ domain (<a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092102">BBa_K1092102</a>), which links  this part together with the PDZ domain-Voltage sensor peptide (<a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092008">BBa_K1092008</a>).<br />
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     <br />
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   </p>
   </p>
     <h4>ssDsbA-PDZ Domain-Voltage Switch (Protein Domain <a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092008">BBa_K1092008</a>)</h4>
     <h4>ssDsbA-PDZ Domain-Voltage Switch (Protein Domain <a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092008">BBa_K1092008</a>)</h4>
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   <p align="center"><img src="https://static.igem.org/mediawiki/2013/a/a8/Sh4.JPG"></p>
   <p>It is a part of the voltage switch, which consist of the PDZ domain (<a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092102">BBa_K1092102</a>) and the  voltage sensor peptide (<a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092006">BBa_K1092006</a>). PDZ Domain  can form dimers with PDZ Ligand (<a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092101">BBa_K1092101</a>), which links  this part together with the PDZ ligand-Voltage sensor peptide (<a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092007">BBa_K1092007</a>).<br />
   <p>It is a part of the voltage switch, which consist of the PDZ domain (<a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092102">BBa_K1092102</a>) and the  voltage sensor peptide (<a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092006">BBa_K1092006</a>). PDZ Domain  can form dimers with PDZ Ligand (<a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092101">BBa_K1092101</a>), which links  this part together with the PDZ ligand-Voltage sensor peptide (<a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092007">BBa_K1092007</a>).<br />
   </p>
   </p>
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<p>&nbsp;</p>
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  <h3>T7  – RBS – Voltage Sensor Peptide (Device <a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092013">BBa_K1092013</a>)</h3>
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  <p align="center"><img src="https://static.igem.org/mediawiki/2013/7/78/20131.png"><img src="https://static.igem.org/mediawiki/2013/7/74/20132.png"></p>
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<p>It is a protein domain found in a transmembrane potassium ion channel from Aeropyrum pernix, which is highly voltage dependent. It contain positive charges from the 4 arginine residues, and also numerous leucine residues which give it high hydrophobicity. All these together make it an excellent voltage sensing peptide in the transmembrane region. Study has shown that this helical voltage sensor peptide is capable of moving across the membrane under external voltage, which is the key to the voltage dependence of the ion channel (PMID: 12721618). We make use of this feature to construct a novel voltage switch (BBa_K1092007 &amp; BBa_K1092008), which consist of the PDZ domain (BBa_K1092102), PDZ ligand (BBa_K1092101), this voltage sensor peptide itself. To further adapt this voltage sensor peptide into our project, we added in two (GGGGS)3 flexible linker which serve to link other protein domains, and a (PE)5 linker which serve as a rigid rod that project from the sensor peptide to elongate it and increase the separation distance.</p>
   <p>&nbsp;</p>
   <p>&nbsp;</p>
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  <h4>T7 - RBS - PDZ Domain - Voltage Sensor – RFP C Terminus (Protein Domain <a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092024">BBa_K1092024</a>)</h4>
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    <p align="center"><img src="https://static.igem.org/mediawiki/2013/4/40/S2.png"></p>
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    <p>This is one protein fragment of the voltage switch system. PDZ domain is  a part of dimer, linked to voltage sensor peptide (BBa_K1092006).</p>
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  <p>&nbsp;</p>
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   <h4>RFP N terminus (Protein Domain <a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092105">BBa_K1092105</a>)</h4>
   <h4>RFP N terminus (Protein Domain <a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092105">BBa_K1092105</a>)</h4>
   
   
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   <p>An RFP fragment adapted from the RFP part <a href="http://partsregistry.org/Part:BBa_E1010">BBa_E1010</a>, and its  counterpart is the RFP N terminus (<a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092105">BBa_K1092105</a>). It is  adapted to be expressed downstream of a fusion protein such as our Voltage  switch parts (<a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092023">BBa_K1092023</a> &amp; <a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092024">BBa_K1092024</a>). It is the C  terminal of the RFP, and is capable of refolding with it&rsquo;s counter part N  terminal as demonstrated by the NYMU-Taipei 2010 iGEM  team through the Biomolecular Fluorescent complementation (BiFC) to give  fluorescent (<a href="http://partsregistry.org/wiki/index.php?title=Part:BBa_K411103">BBa_K411103</a> &amp; <a href="http://partsregistry.org/wiki/index.php?title=Part:BBa_K411104">BBa_K411104</a>). We adapt  this protein to be the downstream of our voltage switch because we want it to  become our effector of the switch. </p>
   <p>An RFP fragment adapted from the RFP part <a href="http://partsregistry.org/Part:BBa_E1010">BBa_E1010</a>, and its  counterpart is the RFP N terminus (<a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092105">BBa_K1092105</a>). It is  adapted to be expressed downstream of a fusion protein such as our Voltage  switch parts (<a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092023">BBa_K1092023</a> &amp; <a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1092024">BBa_K1092024</a>). It is the C  terminal of the RFP, and is capable of refolding with it&rsquo;s counter part N  terminal as demonstrated by the NYMU-Taipei 2010 iGEM  team through the Biomolecular Fluorescent complementation (BiFC) to give  fluorescent (<a href="http://partsregistry.org/wiki/index.php?title=Part:BBa_K411103">BBa_K411103</a> &amp; <a href="http://partsregistry.org/wiki/index.php?title=Part:BBa_K411104">BBa_K411104</a>). We adapt  this protein to be the downstream of our voltage switch because we want it to  become our effector of the switch. </p>
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    <p>&nbsp;</p>
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Latest revision as of 02:28, 29 October 2013

iGEM CUHK