Difference between revisions of "Part:BBa K5477014"
 
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LexA-mERα Chimeric activator with LexA DNA binding domain fused with the mutated ligand-binding domain of Estrogen receptor alpha
 
LexA-mERα Chimeric activator with LexA DNA binding domain fused with the mutated ligand-binding domain of Estrogen receptor alpha
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Full Description: The LexA-mERα (LexA-mutant Estrogen Receptor Alpha) encoding for the chimeric activator where the ligand-binding domain (LBD) of the Estrogen Receptor alpha (ERα) has been mutated. Like the wild-type version, it combines the DNA-binding domain (DBD) of the LexA with the mutated LBD of ERα, designed to bind specific ligands—in this case, modified to potentially alter ligand specificity or binding efficiency. The LexA DBD remains responsible for binding to LexA operator sequences (Lex6Op) in the promoter region, while the mutant LBD (mERα) is responsible for recognizing and responding to specific estrogen-like molecules or analogs, such as bisphenol A (BPA), with altered dynamics compared to the wild-type ERα.
+
Full Description: The LexA-mERα (LexA-mutant Estrogen Receptor Alpha) encoding for the chimeric activator where the ligand-binding domain (LBD) of the Estrogen Receptor alpha (ERα) has been mutated. Like the wild-type version, it combines the DNA-binding domain (DBD) of the LexA with the mutated LBD of ERα, designed to bind specific ligands—in this case, modified to potentially alter ligand specificity or binding efficiency. The LexA DBD remains responsible for binding to LexA operator sequences (Lex6Op) in the promoter region, while the mutant LBD (mERα) is responsible for recognizing and responding to specific estrogen-like molecules or analogs, such as bisphenol A (BPA), with altered dynamics compared to the wild-type ERα (1) (2) (3) (4).
  
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<div style="text-align: center;">
 
https://static.igem.wiki/teams/5477/table-bpa-estrogen.png
 
https://static.igem.wiki/teams/5477/table-bpa-estrogen.png
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</div>
 +
  
 
<p>
 
<p>
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The figure shows the limits of detection (LOD) and half-maximal effective concentrations (EC<sub>50</sub>) for both wild-type (WT) and various mutant versions of the ERα receptor in response to bisphenol A (BPA) and 17β-estradiol. The mutant P4E C8 demonstrates high sensitivity to BPA, with a low EC<sub>50</sub> (1.1 x 10<sup>-6</sup> M) and LOD (2.7 x 10<sup>-7</sup> M), making it highly responsive to BPA while showing minimal sensitivity to 17β-estradiol (EC<sub>50</sub> > 900 nM). This mutant is ideal for specific BPA detection.
+
The figure shows the limits of detection (LOD) and half-maximal effective concentrations (EC<sub>50</sub>) for both wild-type (WT) and various mutant versions of the ERα receptor in response to bisphenol A (BPA) and 17β-estradiol. The mutant P4E C8 demonstrates high sensitivity to BPA, with a low EC<sub>50</sub> (1.1 x 10<sup>-6</sup> M) and LOD (2.7 x 10<sup>-7</sup> M), making it highly responsive to BPA while showing minimal sensitivity to 17β-estradiol (EC<sub>50</sub> > 900 nM). This mutant is ideal for specific BPA detection (3).
 
</p>
 
</p>
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https://static.igem.wiki/teams/5477/era-mutants-alignment.png  
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 +
<div style="text-align: center;">
 +
https://static.igem.wiki/teams/5477/era-alignments.png
 +
</div>
 +
 
 
<p>
 
<p>
 
From the figure above which is from the paper of Rajasärkkä et al. 2011, the mutant P4E C8 mutant version of ERα demonstrates a high sensitivity to bisphenol A (BPA) with a low EC<sub>50</sub> value of 1.1 x 10<sup>-6</sup> M, indicating it is more responsive to BPA compared to other variants. Additionally, it shows low sensitivity to 17β-estradiol, with an EC<sub>50</sub> > 900 nM, demonstrating that this mutant is effective for detecting BPA while minimizing interaction with its native ligand, estradiol.
 
From the figure above which is from the paper of Rajasärkkä et al. 2011, the mutant P4E C8 mutant version of ERα demonstrates a high sensitivity to bisphenol A (BPA) with a low EC<sub>50</sub> value of 1.1 x 10<sup>-6</sup> M, indicating it is more responsive to BPA compared to other variants. Additionally, it shows low sensitivity to 17β-estradiol, with an EC<sub>50</sub> > 900 nM, demonstrating that this mutant is effective for detecting BPA while minimizing interaction with its native ligand, estradiol.
 
</p>
 
</p>
 +
 +
In our system, this part was used as a receptor module to detect estrogenic compounds like BPA that have endocrine disrupting effects in breast milk (1) (2) (3). For more details about how this part was used in our devices, visit the following links:
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 +
 +
===Results from Drylab===
  
  
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===References===
 
===References===
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Rajasärkkä, J., Hakkila, K. and Virta, M. (2011), Developing a compound-specific receptor for bisphenol a by directed evolution of human estrogen receptor ᆇ. Biotechnol. Bioeng., 108: 2526-2534. https://doi.org/10.1002/bit.23214
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1. Çiftçi S, Yalçın SS, Samur G. Bisphenol A Exposure in Exclusively Breastfed Infants and Lactating Women: An Observational Cross-sectional Study. J Clin Res Pediatr Endocrinol. 2021 Nov 25;13(4):375-383. doi: 10.4274/jcrpe.galenos.2020.2021.0305. Epub 2021 Mar 22. PMID: 33749218; PMCID: PMC8638632.
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 +
2. Park, Choa & Song, Heewon & Choi, Junyeong & Sim, Seunghye & Kojima, Hiroyuki & Park, Joonwoo & Iida, Mitsuru & Lee, Youngjoo. (2020). The mixture effects of bisphenol derivatives on estrogen receptor and androgen receptor. Environmental Pollution. 260. 114036. 10.1016/j.envpol.2020.114036.
 +
 
 +
3. Rajasärkkä, J., Hakkila, K. and Virta, M. (2011), Developing a compound-specific receptor for bisphenol a by directed evolution of human estrogen receptor ᆇ. Biotechnol. Bioeng., 108: 2526-2534. https://doi.org/10.1002/bit.23214
 
Zhou, T., Liang, Z. & Marchisio, M.A. Engineering a two-gene system to operate as a highly sensitive biosensor or a sharp switch upon induction with β-estradiol. Sci Rep 12, 21791 (2022). https://doi.org/10.1038/s41598-022-26195-x
 
Zhou, T., Liang, Z. & Marchisio, M.A. Engineering a two-gene system to operate as a highly sensitive biosensor or a sharp switch upon induction with β-estradiol. Sci Rep 12, 21791 (2022). https://doi.org/10.1038/s41598-022-26195-x
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 +
4. Zhou, T., Liang, Z. & Marchisio, M.A. Engineering a two-gene system to operate as a highly sensitive biosensor or a sharp switch upon induction with β-estradiol. Sci Rep 12, 21791 (2022). https://doi.org/10.1038/s41598-022-26195-x

Latest revision as of 16:52, 26 September 2024


LexA-mERα Chimeric activator with LexA DNA binding domain fused with the mutant ERa

LexA-mERα Chimeric activator with LexA DNA binding domain fused with the mutated ligand-binding domain of Estrogen receptor alpha Full Description: The LexA-mERα (LexA-mutant Estrogen Receptor Alpha) encoding for the chimeric activator where the ligand-binding domain (LBD) of the Estrogen Receptor alpha (ERα) has been mutated. Like the wild-type version, it combines the DNA-binding domain (DBD) of the LexA with the mutated LBD of ERα, designed to bind specific ligands—in this case, modified to potentially alter ligand specificity or binding efficiency. The LexA DBD remains responsible for binding to LexA operator sequences (Lex6Op) in the promoter region, while the mutant LBD (mERα) is responsible for recognizing and responding to specific estrogen-like molecules or analogs, such as bisphenol A (BPA), with altered dynamics compared to the wild-type ERα (1) (2) (3) (4).

table-bpa-estrogen.png


The figure shows the limits of detection (LOD) and half-maximal effective concentrations (EC50) for both wild-type (WT) and various mutant versions of the ERα receptor in response to bisphenol A (BPA) and 17β-estradiol. The mutant P4E C8 demonstrates high sensitivity to BPA, with a low EC50 (1.1 x 10-6 M) and LOD (2.7 x 10-7 M), making it highly responsive to BPA while showing minimal sensitivity to 17β-estradiol (EC50 > 900 nM). This mutant is ideal for specific BPA detection (3).

era-alignments.png

From the figure above which is from the paper of Rajasärkkä et al. 2011, the mutant P4E C8 mutant version of ERα demonstrates a high sensitivity to bisphenol A (BPA) with a low EC50 value of 1.1 x 10-6 M, indicating it is more responsive to BPA compared to other variants. Additionally, it shows low sensitivity to 17β-estradiol, with an EC50 > 900 nM, demonstrating that this mutant is effective for detecting BPA while minimizing interaction with its native ligand, estradiol.

In our system, this part was used as a receptor module to detect estrogenic compounds like BPA that have endocrine disrupting effects in breast milk (1) (2) (3). For more details about how this part was used in our devices, visit the following links:


Results from Drylab

Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal XbaI site found at 900
    Illegal PstI site found at 1084
    Illegal PstI site found at 1255
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal PstI site found at 1084
    Illegal PstI site found at 1255
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 1034
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal XbaI site found at 900
    Illegal PstI site found at 1084
    Illegal PstI site found at 1255
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal XbaI site found at 900
    Illegal PstI site found at 1084
    Illegal PstI site found at 1255
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 934


References

1. Çiftçi S, Yalçın SS, Samur G. Bisphenol A Exposure in Exclusively Breastfed Infants and Lactating Women: An Observational Cross-sectional Study. J Clin Res Pediatr Endocrinol. 2021 Nov 25;13(4):375-383. doi: 10.4274/jcrpe.galenos.2020.2021.0305. Epub 2021 Mar 22. PMID: 33749218; PMCID: PMC8638632.

2. Park, Choa & Song, Heewon & Choi, Junyeong & Sim, Seunghye & Kojima, Hiroyuki & Park, Joonwoo & Iida, Mitsuru & Lee, Youngjoo. (2020). The mixture effects of bisphenol derivatives on estrogen receptor and androgen receptor. Environmental Pollution. 260. 114036. 10.1016/j.envpol.2020.114036.

3. Rajasärkkä, J., Hakkila, K. and Virta, M. (2011), Developing a compound-specific receptor for bisphenol a by directed evolution of human estrogen receptor ᆇ. Biotechnol. Bioeng., 108: 2526-2534. https://doi.org/10.1002/bit.23214 Zhou, T., Liang, Z. & Marchisio, M.A. Engineering a two-gene system to operate as a highly sensitive biosensor or a sharp switch upon induction with β-estradiol. Sci Rep 12, 21791 (2022). https://doi.org/10.1038/s41598-022-26195-x

4. Zhou, T., Liang, Z. & Marchisio, M.A. Engineering a two-gene system to operate as a highly sensitive biosensor or a sharp switch upon induction with β-estradiol. Sci Rep 12, 21791 (2022). https://doi.org/10.1038/s41598-022-26195-x