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Rational modification of estrogen receptor by combination of computational and experimental analysis.

Ferrero VE, Pedotti M, Chiadò A, Simonelli L, Calzolai L, Varani L, Lettieri T - PLoS ONE (2014)

Bottom Line: In this manuscript, we modulate the binding properties of estrogen receptor protein by rationally modifying the amino acid composition of its ligand binding domain.These predictions were experimentally confirmed by producing single point variants with up to an order of magnitude increased binding affinity towards some estrogen disrupting chemicals and reaching an half maximal inhibitory concentration (IC50) value of 2 nM for the 17α-ethinylestradiol ligand.Due to increased affinity and stability, utilizing such mutated estrogen receptor instead of the wild type as bio-recognition element would be beneficial in an assay or biosensor.

View Article: PubMed Central - PubMed

Affiliation: European Commission - Directorate General - Joint Research Centre, Institute for Environment and Sustainability, Ispra, Varese, Italy.

ABSTRACT
In this manuscript, we modulate the binding properties of estrogen receptor protein by rationally modifying the amino acid composition of its ligand binding domain. By combining sequence alignment and structural analysis of known estrogen receptor-ligand complexes with computational analysis, we were able to predict estrogen receptor mutants with altered binding properties. These predictions were experimentally confirmed by producing single point variants with up to an order of magnitude increased binding affinity towards some estrogen disrupting chemicals and reaching an half maximal inhibitory concentration (IC50) value of 2 nM for the 17α-ethinylestradiol ligand. Due to increased affinity and stability, utilizing such mutated estrogen receptor instead of the wild type as bio-recognition element would be beneficial in an assay or biosensor.

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Schematic representation of the structural domains of ER protein.A) A cartoon representation of the three-dimensional structure of the ligand binding domain is shown, as well as its sequence. Residues belonging to the ligand binding pocket are shown in red in both the structure and sequence. Residues highlighted in yellow belong to the histidine tag, residues in light blue encompass the ligand binding domain. B) Degree of conservation for residues of the ligand binding pocket among the analyzed ER sequences. Full bars correspond to 100% conservation.
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pone-0102658-g001: Schematic representation of the structural domains of ER protein.A) A cartoon representation of the three-dimensional structure of the ligand binding domain is shown, as well as its sequence. Residues belonging to the ligand binding pocket are shown in red in both the structure and sequence. Residues highlighted in yellow belong to the histidine tag, residues in light blue encompass the ligand binding domain. B) Degree of conservation for residues of the ligand binding pocket among the analyzed ER sequences. Full bars correspond to 100% conservation.

Mentions: The full-length Estrogen Receptor α (ERα) is a protein of approximately 65 kDa formed by several independent structural domains (Figure 1A). The so-called Ligand Binding Domain (LBD, approximately 25 kDa) is necessary and sufficient to bind either the natural ligand (17β-estradiol) or EDCs.


Rational modification of estrogen receptor by combination of computational and experimental analysis.

Ferrero VE, Pedotti M, Chiadò A, Simonelli L, Calzolai L, Varani L, Lettieri T - PLoS ONE (2014)

Schematic representation of the structural domains of ER protein.A) A cartoon representation of the three-dimensional structure of the ligand binding domain is shown, as well as its sequence. Residues belonging to the ligand binding pocket are shown in red in both the structure and sequence. Residues highlighted in yellow belong to the histidine tag, residues in light blue encompass the ligand binding domain. B) Degree of conservation for residues of the ligand binding pocket among the analyzed ER sequences. Full bars correspond to 100% conservation.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4116177&req=5

pone-0102658-g001: Schematic representation of the structural domains of ER protein.A) A cartoon representation of the three-dimensional structure of the ligand binding domain is shown, as well as its sequence. Residues belonging to the ligand binding pocket are shown in red in both the structure and sequence. Residues highlighted in yellow belong to the histidine tag, residues in light blue encompass the ligand binding domain. B) Degree of conservation for residues of the ligand binding pocket among the analyzed ER sequences. Full bars correspond to 100% conservation.
Mentions: The full-length Estrogen Receptor α (ERα) is a protein of approximately 65 kDa formed by several independent structural domains (Figure 1A). The so-called Ligand Binding Domain (LBD, approximately 25 kDa) is necessary and sufficient to bind either the natural ligand (17β-estradiol) or EDCs.

Bottom Line: In this manuscript, we modulate the binding properties of estrogen receptor protein by rationally modifying the amino acid composition of its ligand binding domain.These predictions were experimentally confirmed by producing single point variants with up to an order of magnitude increased binding affinity towards some estrogen disrupting chemicals and reaching an half maximal inhibitory concentration (IC50) value of 2 nM for the 17α-ethinylestradiol ligand.Due to increased affinity and stability, utilizing such mutated estrogen receptor instead of the wild type as bio-recognition element would be beneficial in an assay or biosensor.

View Article: PubMed Central - PubMed

Affiliation: European Commission - Directorate General - Joint Research Centre, Institute for Environment and Sustainability, Ispra, Varese, Italy.

ABSTRACT
In this manuscript, we modulate the binding properties of estrogen receptor protein by rationally modifying the amino acid composition of its ligand binding domain. By combining sequence alignment and structural analysis of known estrogen receptor-ligand complexes with computational analysis, we were able to predict estrogen receptor mutants with altered binding properties. These predictions were experimentally confirmed by producing single point variants with up to an order of magnitude increased binding affinity towards some estrogen disrupting chemicals and reaching an half maximal inhibitory concentration (IC50) value of 2 nM for the 17α-ethinylestradiol ligand. Due to increased affinity and stability, utilizing such mutated estrogen receptor instead of the wild type as bio-recognition element would be beneficial in an assay or biosensor.

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