Difference between revisions of "Part:BBa K4698007"
 
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A protein degradation tag (degron) is attached to the C-terminus of GFP to promote protein degradation.
 
A protein degradation tag (degron) is attached to the C-terminus of GFP to promote protein degradation.
  
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===Usage and Biology===
 
===Usage and Biology===
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While reviewing CRISPR-related materials, we discovered a particularly interesting evolutionary system- phage-assisted continuous evolution (PACE)[1,2], which can enable continuous directed evolution of gene-coding molecules that can be associated with protein production in <i>E. coli</i>. Then, we found a protein degradation tag-degron[3], that enables rapid hydrolysis of proteins with little functionality, which allows degron to be used in a number of protein co-expression screening systems.
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<p>While reviewing CRISPR-related materials, we discovered a particularly interesting evolutionary system- phage-assisted continuous evolution (PACE)[1,2], which can enable continuous directed evolution of gene-coding molecules that can be associated with protein production in <i>E. coli</i>. Then, we found a protein degradation tag-degron[3], that enables rapid hydrolysis of proteins with little functionality, which allows degron to be used in a number of protein co-expression screening systems.</p>
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<p>We expressed GFP in fusion with degron and tested the degradation efficiency of degron (Fig. 1). The results showed that the GFP-degron fusion protein was almost non-fluorescent, much less than GFP. this demonstrated the extremely strong degradation activity of degron.</p>
 
<img src="https://static.igem.wiki/teams/4698/wiki/improved-part-gfp.png" class= "center" style="width:500px">
 
<img src="https://static.igem.wiki/teams/4698/wiki/improved-part-gfp.png" class= "center" style="width:500px">
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Figure 1, Testing the protein degradation efficiency of degron. blank, LB medium. NT, E.coli without GFP expression plasmid, n=3.
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<p>Figure 1, Testing the protein degradation efficiency of degron. blank, LB medium. NT, <i>E.coli</i> without GFP expression plasmid, <i>n</i>=3.</p>
  
  
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2. Miller, S. M., Wang, T. & Liu, D. R. Phage-assisted continuous and non-continuous evolution. <i>Nature Protocols</i> 15, 4101-4127 (2020).
 
2. Miller, S. M., Wang, T. & Liu, D. R. Phage-assisted continuous and non-continuous evolution. <i>Nature Protocols</i> 15, 4101-4127 (2020).
 
3. Karzai, A. W., Roche, E. D. & Sauer, R. T. The SsrA-SmpB system for protein tagging, directed degradation and ribosome rescue. <i>Nat Struct Biol</i> 7, 449-455 (2000).
 
3. Karzai, A. W., Roche, E. D. & Sauer, R. T. The SsrA-SmpB system for protein tagging, directed degradation and ribosome rescue. <i>Nat Struct Biol</i> 7, 449-455 (2000).
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<span class='h3bb'>Sequence and Features</span>
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<partinfo>BBa_K4698008 SequenceAndFeatures</partinfo>
  
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===Functional Parameters===
 
===Functional Parameters===
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<partinfo>BBa_K4698007 parameters</partinfo>
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<partinfo>BBa_K4698008 parameters</partinfo>
 
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Latest revision as of 15:26, 11 October 2023


GFP with a protein degradation tag attached

A protein degradation tag (degron) is attached to the C-terminus of GFP to promote protein degradation.

Usage and Biology

While reviewing CRISPR-related materials, we discovered a particularly interesting evolutionary system- phage-assisted continuous evolution (PACE)[1,2], which can enable continuous directed evolution of gene-coding molecules that can be associated with protein production in E. coli. Then, we found a protein degradation tag-degron[3], that enables rapid hydrolysis of proteins with little functionality, which allows degron to be used in a number of protein co-expression screening systems.

We expressed GFP in fusion with degron and tested the degradation efficiency of degron (Fig. 1). The results showed that the GFP-degron fusion protein was almost non-fluorescent, much less than GFP. this demonstrated the extremely strong degradation activity of degron.

Figure 1, Testing the protein degradation efficiency of degron. blank, LB medium. NT, E.coli without GFP expression plasmid, n=3.

Refrence

1. Esvelt, K. M., Carlson, J. C. & Liu, D. R. A system for the continuous directed evolution of biomolecules. Nature 472, 499-503 (2011). 2. Miller, S. M., Wang, T. & Liu, D. R. Phage-assisted continuous and non-continuous evolution. Nature Protocols 15, 4101-4127 (2020). 3. Karzai, A. W., Roche, E. D. & Sauer, R. T. The SsrA-SmpB system for protein tagging, directed degradation and ribosome rescue. Nat Struct Biol 7, 449-455 (2000).

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]