Difference between revisions of "Part:BBa K2317006"

 
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DmpR is a protein that can be used as a cholophenol sensor.
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DmpR is a protein that can be used as a cholophenol sensor.<br>
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<br>
 
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===Usage and Biology===
 
===Usage and Biology===
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<partinfo>BBa_K2317006 parameters</partinfo>
 
<partinfo>BBa_K2317006 parameters</partinfo>
 
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<head></head>
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<body>
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<h3>Description</h3>
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<p>DmpR, a protein coded by Pseudomonas sp. Strain CF600 <i>dmpR</i> gene [1-2], shows its ability to response to simple phenols. And in our circuit, when detecting the presence of an inducing phenol, a productive association between the sensor domain and a phenolic molecule will lead to DmpR undergoing a conformational change that results in a polymerase-activating form of the protein, promoting the expression of gene downstream promoter Po[3,4].</p>
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<br/>
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<div class="pic_box center">
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<img src="https://static.igem.org/mediawiki/2017/e/e4/T--Jilin_China--design002.png" width="60%" /><br />
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Figure 1. The mechanism of DmpR sensor.
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</div>
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<p>However, on the account of some research on the mutation DmpR finding the mutated one shows their more sensitive response and wider substrates range[5,6], so we selected one mutated DmpR. The mutant type DmpR we use has 5 sites of nucleotides mutation[7] of which two are nonsense mutation and others lead to two amino acids change. </p>
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<div class="pic_box center">
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<img src="https://static.igem.org/mediawiki/2017/c/c5/T--Jilin_China--composite_parts02.png"  />
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<img src="https://static.igem.org/mediawiki/2017/0/0a/T--Jilin_China--composite_parts03.png"  />
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<img src="https://static.igem.org/mediawiki/2017/5/57/T--Jilin_China--composite_parts04.png"  /><br />
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Figure 2. Mutant type DmpR and wild type DmpR
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</div><br/>
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<p>What's more, Peking University has registered a part (<a href="https://parts.igem.org/Part:BBa_K1031211">BBa_K1031211</a>) including wild-type DmpR in Parts Registry, so we made some comparisons on the substrates between our DmpR and Peking’s DmpR. And the result is as follow.</p>
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<br/>
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<div class="pic_box center">
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<img src="https://static.igem.org/mediawiki/2017/7/7c/T--Jilin_China--improvement04.png" width="60%" /><br />
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Figure 3. DmpR response sensitivity toward different phenolic compounds.
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</div><br/>
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<p>We used the ratio of promotors response and mock response to the same phenolic compounds to compare the differences between wild type DmpR and mutant type DmpR.</p>
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<p>The phenolic compounds we chose included: phenol, 2-CP, 4-CP, pyrocatechol,2,4-DCP</p>
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<br/>
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<p>According to the result we get, it turned out that wild type DmpR can response to phenol, 2-CP, pyrocatechol. Compared with wild type DmpR in BBa_K1031211, our mutant type DmpR showed better response to phenol and pyrocatechol, which means our mutant type DmpR is more efficient.</p>
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<br/>
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<p>For more information, <a href="http://2017.igem.org/Team:Jilin_China/Improve">please visit...</a></p>
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<br/>
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<h3>Reference</h3>
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<p>[1] Shingler, V., M. Bartilson, and T. Moore. Cloning and nucleotide sequence of the gene encoding the positive regulator (DmpR) of the phenol catabolic pathway encoded by pVI150 and identification of DmpR as a member of the NtrC family of transcriptional activators. J. Bacteriol. 1993,175: 1596–1604.</p>
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<p>[2] Shingler, V., and T. Moore. Sensing of aromatic compounds by the DmpR transcriptional activator of phenol-catabolizing Pseudomonas sp. strain CF600. J. Bacteriol. 1994, 176:1555–1560.</p>
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<p>[3] Ng, L., E. O’Neil, and V. Shingler. Genetic evidence for interdomain regulation of the phenol-responsive s54-dependent activator DmpR. J. Biol. Chem. 1996, 271:17281–17286.</p>
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<p>[4] Shingler, V., and H. Pavel. Direct regulation of the ATPase activity of the transcriptional activator DmpR by arromatic compounds. Mol. Microbiol.1995, 17:505–513.</p>
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<p>[5] V. L. Campos,J. Veas, C. A. Zaror, and M. A. Mondaca1. Monitoring Phenolic Compounds During Biological Treatment of Kraft Pulp Mill Effluent Using Bacterial Biosensors. Bull. Environ. Contam. Toxicol. 2006, 77:383–390 </p>
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<p>[6]V. L. Campos,C. A. Zaror, and M. A. Mondaca.Detection of chlorinated phenols in kraft pulp bleaching effluents using DmpR mutant strains. Bull. Environ. Contam. Toxicol. 2004, 73:666–673 </p>
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<p>[7]Arlene A. Wise and Cheryl R. Kuske. Generation of novel bacterial regulatory proteins that detect priority pollutant phenols. Appl. Environ. Microb. 2000, 163-169</p>
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</body>
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</html>

Revision as of 11:42, 1 November 2017

DmpR

DmpR is a protein that can be used as a cholophenol sensor.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 86
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 564
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 249
    Illegal BsaI.rc site found at 790
    Illegal SapI.rc site found at 1447


Description

DmpR, a protein coded by Pseudomonas sp. Strain CF600 dmpR gene [1-2], shows its ability to response to simple phenols. And in our circuit, when detecting the presence of an inducing phenol, a productive association between the sensor domain and a phenolic molecule will lead to DmpR undergoing a conformational change that results in a polymerase-activating form of the protein, promoting the expression of gene downstream promoter Po[3,4].



Figure 1. The mechanism of DmpR sensor.

However, on the account of some research on the mutation DmpR finding the mutated one shows their more sensitive response and wider substrates range[5,6], so we selected one mutated DmpR. The mutant type DmpR we use has 5 sites of nucleotides mutation[7] of which two are nonsense mutation and others lead to two amino acids change. 


Figure 2. Mutant type DmpR and wild type DmpR

What's more, Peking University has registered a part (BBa_K1031211) including wild-type DmpR in Parts Registry, so we made some comparisons on the substrates between our DmpR and Peking’s DmpR. And the result is as follow.



Figure 3. DmpR response sensitivity toward different phenolic compounds.

We used the ratio of promotors response and mock response to the same phenolic compounds to compare the differences between wild type DmpR and mutant type DmpR.

The phenolic compounds we chose included: phenol, 2-CP, 4-CP, pyrocatechol,2,4-DCP


According to the result we get, it turned out that wild type DmpR can response to phenol, 2-CP, pyrocatechol. Compared with wild type DmpR in BBa_K1031211, our mutant type DmpR showed better response to phenol and pyrocatechol, which means our mutant type DmpR is more efficient.


For more information, please visit...


Reference

[1] Shingler, V., M. Bartilson, and T. Moore. Cloning and nucleotide sequence of the gene encoding the positive regulator (DmpR) of the phenol catabolic pathway encoded by pVI150 and identification of DmpR as a member of the NtrC family of transcriptional activators. J. Bacteriol. 1993,175: 1596–1604.

[2] Shingler, V., and T. Moore. Sensing of aromatic compounds by the DmpR transcriptional activator of phenol-catabolizing Pseudomonas sp. strain CF600. J. Bacteriol. 1994, 176:1555–1560.

[3] Ng, L., E. O’Neil, and V. Shingler. Genetic evidence for interdomain regulation of the phenol-responsive s54-dependent activator DmpR. J. Biol. Chem. 1996, 271:17281–17286.

[4] Shingler, V., and H. Pavel. Direct regulation of the ATPase activity of the transcriptional activator DmpR by arromatic compounds. Mol. Microbiol.1995, 17:505–513.

[5] V. L. Campos,J. Veas, C. A. Zaror, and M. A. Mondaca1. Monitoring Phenolic Compounds During Biological Treatment of Kraft Pulp Mill Effluent Using Bacterial Biosensors. Bull. Environ. Contam. Toxicol. 2006, 77:383–390

[6]V. L. Campos,C. A. Zaror, and M. A. Mondaca.Detection of chlorinated phenols in kraft pulp bleaching effluents using DmpR mutant strains. Bull. Environ. Contam. Toxicol. 2004, 73:666–673

[7]Arlene A. Wise and Cheryl R. Kuske. Generation of novel bacterial regulatory proteins that detect priority pollutant phenols. Appl. Environ. Microb. 2000, 163-169