Difference between revisions of "Team:KAIT Japan/Description"

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<h2> Project Description </h2>
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<p>Tell us about your project, describe what moves you and why this is something important for your team.</p>
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<li> A clear and concise description of your project.</li>
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<h4>Advice on writing your Project Description</h4>
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We encourage you to put up a lot of information and content on your wiki, but we also encourage you to include summaries as much as possible. If you think of the sections in your project description as the sections in a publication, you should try to be consist, accurate and unambiguous in your achievements.
 
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<h3><a href="https://2015.igem.org/Team:KAIT_Japan"><img src="https://static.igem.org/mediawiki/2015/thumb/5/54/%E3%83%AD%E3%82%B4%E3%83%9E%E3%83%BC%E3%82%AF.jpg/800px-%E3%83%AD%E3%82%B4%E3%83%9E%E3%83%BC%E3%82%AF.jpg"  width="90px" height="70px"></a><FONT size="11">Project</FONT></h3>
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<br>&nbsp;
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<br><font size="6">"Control protein activity by light"</font><br><br>
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&nbsp;&nbsp;In this year, we use the fluorescent protein, “Dronpa“, to make protein regulation system. Dronpa has photochromism, Dronpa able to change its state bright from dark or dark from bright.  Dronpa normally absorbs at 503 nm and emits green fluorescence, strong irradiation at 488 nm can convert this protein to a nonfluorescent state that absorbs at 390 nm. The protein can then be switched back to the original emissive state with minimal irradiation at 405 nm.<br>
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&nbsp;&nbsp;We use Dronpa145N which is Lys145Asn mutant of Dronpa. At the bright state, this protein forms tetramer. While at dark state, Dronpa145N form monomer. The ensemble absorption and emission properties of Dronpa145N are highly similar to those of Dronpa. We utilize this conformation change for Light-dependent regulation of flexibility in a protein.<br>
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<img src="https://static.igem.org/mediawiki/2015/5/59/%EF%BC%A6%EF%BC%91.png"><br>
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<font size="3">[Figure 1]:The tetrameric structure of Dronpa145N.</font><br>
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<br>
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<br>
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<img src="https://static.igem.org/mediawiki/2015/1/11/Tiba_syuusei.PNG"><br>
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<font size="3">[Figure 2]:Overview of photochromism of Dronpa145N.</font><br>
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<br>
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<br>
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&nbsp;&nbsp;Lys145Asn mutation effect the oligomerization-interface of this protein. In Dronpa, a high degrees of freedom of Lys145 make repulsion between one protomer and another protomer. But in Dronpa145N, it’s repulsion is fewer than Dronpa, because Asn145 has fewer degrees of freedom compared with Lys145’s it. Therefor Dronpa145N is able to tetramerization.<br>
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<br>
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<img src="https://static.igem.org/mediawiki/2015/f/f3/%EF%BC%A6%EF%BC%94.png2.png"><br>
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<font size="3">[Figure 3]:Overview of mutation of Dronpa145N from Dronpa.</font><br>
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<br>
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<br>
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&nbsp;&nbsp;We fuse target protein between Dronpa145N. When Dronpa145N is illuminated at 405nm, both side of Dronpa145N form dimer and conformation of target protein is bent. While when Dronpa145N is illuminated at 488nm, dimer change monomer and target protein form original conformation. As a result, the target protein’s activity is reduced.<br>
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<br>
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<img src="https://static.igem.org/mediawiki/2015/9/94/%EF%BC%A6%EF%BC%93.png2.png"><br>
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<font size="3">[Figure 4]:(a)Overview of the AC interfaces of Dronpa145N. Two cylinder show β-barrel and grey circles are AB interfaces. (b)Closer view of the details of the AC interfaces showing parts of the β-strands. β-strands are shown as black lines and relevant residues are shown in Y. Black two headed arrows indicate electrostatic interaction. Orange two headed arrows indicate hydrophobic interaction. Color code: polar uncharged residues, green; negative charged residues, red; positive charged residues, blue.</font><br>
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<br>
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<br>
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<li><a href="https://2015.igem.org/Team:KAIT_Japan/Results"><font size="5">Results</font></a></li>
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<li><a href="https://2015.igem.org/Team:KAIT_Japan/contribution"><font size="5">Contribution</font></a></li>
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<br>
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<br>
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----Reference----<br>
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1) Ngan Nguyen Bich. et al.“Structural basis for the influence of a single mutation K145N on the oligomerization and photoswiching rate of Dronpa” Acta Cryst,D68,2012,1653-1659<br>
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<br>
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2) Xin X.Zhou and Michael Z. Lin.”Photoswitchable Fluorescent Proteins: Ten Years of Colorful Chemistry and Exciting Applications”Curr Opin Chem Biol,Aug;17(4),2013,682-690<br>
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<br>
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3) Hideaki Mizuno.et al.”Light-dependent regulation of structural flexibility in a photohromic fluorescent protein”Proc Natl Acad Sci U S A,105(27),2008,927-9232<br>
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<br>
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4) XIn X. Zhou. et al.”Optical control of protein activeity by fluorescent protein comains”Science,338(6108),2012,810-814 <br>
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<font size="9" face="Freestyle Script">---Link list---</font>
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<br>
 
<ul>
 
<ul>
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<li><a href="https://2014.igem.org/Team:Imperial/Project"> Imperial</a></li>
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<li><a href="https://2015.igem.org/Team:KAIT_Japan/Attributions">Attributions</a></li>
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<li><a href="https://2014.igem.org/Team:UC_Davis/Project_Overview"> UC Davis</a></li>
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<li><a href="https://2015.igem.org/Team:KAIT_Japan/Team">Team</a></li>
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<li><a href="https://2014.igem.org/Team:SYSU-Software/Overview">SYSU Software</a></li>
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<li><a href="https://2015.igem.org/Team:KAIT_Japan/Results">Results</a></li>
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<li><a href="https://2015.igem.org/Team:KAIT_Japan/Basic_Part">Parts</a></li>
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<li><a href="https://2015.igem.org/Team:KAIT_Japan/Protocol">Protocol</a></li>
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<li><a href="https://2015.igem.org/Team:KAIT_Japan/Notebook">Lab note</a></li>
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<li><a href="https://2015.igem.org/Team:KAIT_Japan/Safety">Safety</a></li>
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<li><a href="https://2015.igem.org/Team:KAIT_Japan/Practices">Human Practice</a></li>
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<li><a href="https://2015.igem.org/Team:KAIT_Japan/Judge">Achievement</a></li>
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<li><a href="https://2015.igem.org/Team:KAIT_Japan/contribution">Contribution</a></li>
 
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<font size="9" face="Freestyle Script">---Sponsor---</font>
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<a href="http://www.cosmobio.co.jp/index_e.asp"><img src="https://static.igem.org/mediawiki/2015/6/62/Cosumo_bio.png" width="180px"></a>
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<br>
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<a href="http://www.kait.jp/yumepro/"><img src="https://static.igem.org/mediawiki/2015/0/07/Yumepuro.png" width="180px"></a>
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<br>
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<br>
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<a href="http://www.promega.co.jp/"><img src="https://static.igem.org/mediawiki/2015/f/f6/Puromega.png" width="180px"></a>
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<br>
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<a href="https://lne.st/"><img src="https://static.igem.org/mediawiki/2015/3/3e/Ribane.png" width="180px"></a>
  
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Latest revision as of 15:23, 18 September 2015

Project


 
"Control protein activity by light"

  In this year, we use the fluorescent protein, “Dronpa“, to make protein regulation system. Dronpa has photochromism, Dronpa able to change its state bright from dark or dark from bright. Dronpa normally absorbs at 503 nm and emits green fluorescence, strong irradiation at 488 nm can convert this protein to a nonfluorescent state that absorbs at 390 nm. The protein can then be switched back to the original emissive state with minimal irradiation at 405 nm.
  We use Dronpa145N which is Lys145Asn mutant of Dronpa. At the bright state, this protein forms tetramer. While at dark state, Dronpa145N form monomer. The ensemble absorption and emission properties of Dronpa145N are highly similar to those of Dronpa. We utilize this conformation change for Light-dependent regulation of flexibility in a protein.

[Figure 1]:The tetrameric structure of Dronpa145N.



[Figure 2]:Overview of photochromism of Dronpa145N.


  Lys145Asn mutation effect the oligomerization-interface of this protein. In Dronpa, a high degrees of freedom of Lys145 make repulsion between one protomer and another protomer. But in Dronpa145N, it’s repulsion is fewer than Dronpa, because Asn145 has fewer degrees of freedom compared with Lys145’s it. Therefor Dronpa145N is able to tetramerization.


[Figure 3]:Overview of mutation of Dronpa145N from Dronpa.


  We fuse target protein between Dronpa145N. When Dronpa145N is illuminated at 405nm, both side of Dronpa145N form dimer and conformation of target protein is bent. While when Dronpa145N is illuminated at 488nm, dimer change monomer and target protein form original conformation. As a result, the target protein’s activity is reduced.


[Figure 4]:(a)Overview of the AC interfaces of Dronpa145N. Two cylinder show β-barrel and grey circles are AB interfaces. (b)Closer view of the details of the AC interfaces showing parts of the β-strands. β-strands are shown as black lines and relevant residues are shown in Y. Black two headed arrows indicate electrostatic interaction. Orange two headed arrows indicate hydrophobic interaction. Color code: polar uncharged residues, green; negative charged residues, red; positive charged residues, blue.


  • Results
  • Contribution


    • ----Reference----
    1) Ngan Nguyen Bich. et al.“Structural basis for the influence of a single mutation K145N on the oligomerization and photoswiching rate of Dronpa” Acta Cryst,D68,2012,1653-1659

    2) Xin X.Zhou and Michael Z. Lin.”Photoswitchable Fluorescent Proteins: Ten Years of Colorful Chemistry and Exciting Applications”Curr Opin Chem Biol,Aug;17(4),2013,682-690

    3) Hideaki Mizuno.et al.”Light-dependent regulation of structural flexibility in a photohromic fluorescent protein”Proc Natl Acad Sci U S A,105(27),2008,927-9232

    4) XIn X. Zhou. et al.”Optical control of protein activeity by fluorescent protein comains”Science,338(6108),2012,810-814