Difference between revisions of "Part:BBa K1980000"
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===Usage and Biology===
 
===Usage and Biology===
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<p> Our project aimed to detect and chelate dietary copper as a treatment for Wilson's Disease, a copper accumulation disorder.
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We decided that the ideal copper chelation protein would have these properties:
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</p>
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<ul class="textLiFont">
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<li>Should be able to bind multiple copper ions per peptide to increase the efficient use of cell resources. </li>
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<li>They should be from the prokaryotic domain because eukaryotic proteins can have expression issues in <i>Escherichia coli.</i> </li>
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<li>As <i> E. coli </i>naturally deals with copper toxicity by binding copper in the periplasm then exporting it, periplasmic proteins may reduce toxicity to the host. </li>
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<p>Using these criteria we found two copper chelators that we thought would be useful. We designed gBlocks, codon optimised for <i>E. coli</i>, containing these parts both alone and linked to super-folder <a data-toggle="popover1" data-trigger="hover" title="Green Fluorescent Protein" data-content="A protein that emits green light when illuminated with blue to UV light">GFP</a>. This form of the fluorescent protein was used because the standard GFP doesn’t fold particularly well in the periplasm where one of our chelators is intended to travel to.
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The proteins, both alone and attached to sfGFP, were ordered with C terminal <a data-toggle="popover1" data-trigger="hover" placement:"top" title="His Tag" data-content="Six sequential Histidine residues that bind to NTA columns"> hexa-histidine tag</a> so we could purify them. A concern was raised that the His tag would also weakly bind copper potentially affecting the results. However we decided that increasing the copper-binding would only improve the proteins' intended function. 
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<span class='h3bb'>Sequence and Features:</span>
 
<span class='h3bb'>Sequence and Features:</span>

Revision as of 19:57, 16 October 2016


TAT Copper Storage Protein 1

Copper Storage protein 1 (Csp1) is a tetrameric copper storage protein found in the periplasm of Methylosinus trichosporium OB3b. We investigated whether this part could act as a copper chelator when expressed in E. coli. We modified the protein by adding a TAT signal peptide from the E. coli enzyme CueO in place of the native TAT sequence and added a C terminal his tag to aid purification.

Sequence and Features:


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]