Part:BBa_K5477013
LexA-ERα - Chimeric activator with LexA DNA binding domain fused with ERα-LBD
The LexA-ERα (LexA-Estrogen Receptor Alpha) fusion protein combines the DNA-binding domain of the bacterial LexA repressor with the ligand-binding domain (LBD) of the Estrogen Receptor alpha (ERα). This engineered protein is designed to utilize the ligand-binding specificity of ERα and the DNA-binding specificity of the LexA operator system. In this fusion, the LexA DBD binds to LexA operator sequences (Lex6Op) , while the ERα LBD allows the regulation of gene expression in response to estrogen or estrogen-like molecules, in this case BPA. When BPA binds to the ERα LBD within this fusion protein, it induces a conformational change that activates the reporter module that has Lex6Op.
This part was based on the study of Zhou et al. 2022. The LexA domain was fused with either the ligand-binding domain of the wild-type ERα, the mutant ERα or the Estrogen-Related Receptor gamma ERRγ. An alignment of the ligand-binding domains of the aforementioned was performed to determine the exact sequence to fuse with LexA to generate a chimeric activator that will bind to the Lex6Op in the reporter module.
Figure 1 - Multiple Sequence Alignment of Estrogen receptors - alpha and beta with Estrogen-related receptor gamma (CLUSTALW)
Figure 2 - 3D Structure Prediction of Estrogen receptor alpha showing residue 281 found in the loop. The curved green line divides the structure and shows the ligand-binding domain of ERα. From residue 281 until the end of the amino acid sequence is fused downstream with LexA DNA binding domain to generate the chimeric activator.
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) (4) (5) (6). For more details about how this part was used in the devices, visit the following links: BBa_K5477043 and BBa_K5477044.
Sequence and Features
- 10INCOMPATIBLE WITH RFC[10]Illegal XbaI site found at 900
Illegal PstI site found at 1084
Illegal PstI site found at 1255 - 12INCOMPATIBLE WITH RFC[12]Illegal PstI site found at 1084
Illegal PstI site found at 1255 - 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 1034
- 23INCOMPATIBLE WITH RFC[23]Illegal XbaI site found at 900
Illegal PstI site found at 1084
Illegal PstI site found at 1255 - 25INCOMPATIBLE WITH RFC[25]Illegal XbaI site found at 900
Illegal PstI site found at 1084
Illegal PstI site found at 1255 - 1000COMPATIBLE WITH RFC[1000]
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. Hafezi SA, Abdel-Rahman WM. The Endocrine Disruptor Bisphenol A (BPA) Exerts a Wide Range of Effects in Carcinogenesis and Response to Therapy. Curr Mol Pharmacol. 2019;12(3):230-238. doi: 10.2174/1874467212666190306164507. PMID: 30848227; PMCID: PMC6864600.
3. Lo, Elena & Piparo, Elena & Siragusa, Lydia & Raymond, Frédéric & Passeri, Giovanna & Cruciani, Gabriele & Schilter, Benoit. (2019). Bisphenol A Binding Promiscuity: A Virtual Journey through the Universe of Proteins 1.
4. 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.
5. 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
6. 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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