Estrogens, their receptors and relevant proteins.
There are three major endogenous estrogens that have estrogenic hormonal activity: estrone (E1), estradiol (E2), and estriol (E3). Estradiol, an estrane, is the most potent and prevalent. Estradiol is an important estrogen steroid hormone in both women and men. It is a typical steroid with core four-ring system (ABCD), composed of 17 carbon atoms. Another estrogen called estetrol (E4) is produced only during pregnancy.
Click here to see the difference between conformations of estrogen receptor α complexed with raloxifene and a corepressor peptide (morph was taken from Gallery of Morphs of the Yale Morph Server).
Structure of estradiol metal chelate and estrogen receptor complex: The basis for designing a new class of SERMs[1].
Selective estrogen receptor modulators, such as estradiol 17-derived metal complexes, have been synthesized as targeted probes for the diagnosis and treatment of breast cancer. The detailed 3D structure of estrogen receptor α ligand-binding domain (ER-LBD) bound with a novel estradiol-derived metal complex, estradiol-pyridinium tetra acetate europium (III) (EPTA-Eu) at 2.6Å resolution was reported (2yat). The residues Glu353, Arg394 and His524 and the conserved water molecule (W1006) form hydrogen bonds with EPTA-Eu. The hydrogen bonds are shown as white dashed lines. Superposition of this structure with the structure of native ligand 17β-estradiol (E2) in the complex of E2/ERα-LBD complex (1ere) reveals that the E2 core of EPTA-Eu overlaps closely with that of E2 itself. The hydrogen bonds network made by additional estrogen receptor residues (e.g. Glu419 of H7 and Glu339 of H3, this depends on subunit), may work together with the E2 17β hydroxyl-His524 hydrogen bond and tighten the neck of the LBP upon binding of the endogenous ligand E2. 4-Hydroxytamoxifen (OHT) is an other selective estrogen receptor modulator. Superposition of EPTA-Eu/ERα-LBD complex on OHT/ERα-LBD complex (3ert) shows that there is similar network of hydrogen bonds in both complexes, except for His524 which does not form hydrogen bond with OHT in the OHT/ERα-LBD complex. Superposition of structures of all these three complexes: E2/ERα-LBD (1ere), OHT/ERα-LBD (3ert) and EPTA-Eu/ERα-LBD shows that they overlap well in the majority portions of the domain, but differ significantly in the region of the 'omega loop'. They display different synergistic reciprocating movements, depending on the specific nature of the ligand bound. The structure of estrogen receptor complexed with EPTA-Eu provides important information pertinent to the design of novel functional ER targeted probes for clinical applications.
ER is a modular protein composed of a ligand binding domain, a DNA binding domain and a transactivation domain. ER is a DNA-binding transcription factor.
ER bound to DNA. The DNA binding domain can be clearly observed in this scene; the highlighted yellow helix in close proximity to the DNA is part of the DNA binding domain. The blue beta sheet close to the yellow DNA binding alpha helix is also part of the DNA binding domain. The transactivation domain forms an alpha helix which is colored in purple. The transactivation domain activates RNA polymerase when the receptor binds to DNA. The ligand binding domain may be observed here with the following scene.
Agonist ferutinine bound ER. The ligand ferutinine (highlighted in pink) is bound by the ligand binding domain, composed of the blue colored alpha helices immediately surrounding the purple ligand. Another view of the ligand binding domain is shown here, with estradiol bound. ER ligand binding domain bound to estradiol.
ER is functional as a ligand-dependent transcription factor. ER responds to both agonist and antagonist ligands and can associate with the nuclear matrix. Differences in the structure of the receptor are observed depending on what ligand ER has bound (if any). Through comparisons of ER bound to agonist and antagonist ligands, some structural components may be highlighted. Agonist estradiol bound er The specific conformation of this tight loop of alpha helices and beta sheets around the ligand shows a complex capable of activating ER's transcription loci. This complex allows for the activation signal that will stimulate normal growth.
Normal growth is stimulated when an agonist bound ER binds DNA. This occurs with the assistance of chaperon proteins. These chaperons are capable of recognizing estrogen receptor ligand complexes. When ER has bound a ligand chaperons facilitate the trans-location of the complex to the nucleus. Eventually the chaperon ligand ER complex will reach specific euchromatin, at which point the chaperons facilitate the ligand ER complex to changes conformation. This conformation will facilitate the estrogen receptor to bind the DNA major groove at specific palindromic sequences. Estradiol is a normal ligand for ER and allows for binding in the major groove of DNA.
If the ligand is an antagonist the transcription factor function of estrogen receptor becomes hindered. Partial Agonist genistein bound ER The conformation of ER bound to the partial agonist genistein has a loop which is not as tight around the ligand as those found on ER with a complete agonist ligand. The ligands themselves take up different amounts of space and have varying interactions within ER. This slight difference effects the ability of the chaperon to be able to bind the receptor ligand complex to the major groove of DNA. There is a noticeable difference in the size of the pure agonist vs partial agonist scenes. Specifically, look at the width of the agonist compared to the partial agonist.
Similar differences may be observed between ER which has bound the partial agonist and complete antagonist ligands. Antagonist tamoxifen bound ER The most drastic difference is noticeable between agonist and antagonist ligands. Compare the agonist scene to the Agonist estradiol bound er. Special attention should be given to the bottom right alpha helices and beta sheets that are pushed out more in the antagonist compared to the agonist bound ER.
Estrogen receptor β (ER-β) is 1 of the 2 isoforms of the estrogen receptor, a ligand-activated transcription factor which regulates the biological effects of the steroid hormone 17 β-estradiol, or estrogen, in both males and females. The complex is a hetero-tetrameric assembly consisting of 4 molecules and a ligand: 2 copies of estrogen receptor β, 2 copies of steroid receptor coactivator-1, and the ligand, Genistein. Once the ligand is bound, the complex recruits the steroid receptor coactivators, which recruit other proteins to form the transcriptional complex for initiation of transcription. This activates expression of reporter genes containing estrogen response elements. Genistein is a phytoestrogen with structural similarity to estrogen which competes for estrogen receptors.
Although estrogen receptor β is widely expressed, it is not the primary estrogen receptor in most tissues. As a result, it has become a target for drug delivery, especially since it is 40x more selective for genistein than the α isoform. This enhanced selectivity may be caused by differences in residues 336 and 373 between the 2 isoforms, allowing ER-β to accommodate more polar substituents in its binding pocket. ER-β differs greatly from ER-α at the N-terminal domains, which can be seen located at opposite ends from the C termini in this rainbow representation. The protein is composed of 3 sections: a modulating N-terminal domain, a DNA-binding domain and a C-terminal ligand-binding domain.
Each ERβ contains several domains with specific functions: an N-terminal domain (NTD), a DNA-binding domain (DBD), a flexible hinge region and a C-terminal Ligand-binding domain (LBD). The complex overall is about 66% helical (10 helices; 160 residues) and 3% β-sheet (2 strands; 9 residues). The NTD is the 1st activation function (AF-1) domain that consists mostly of random coils and a small portion of helices (red) and sheets (green); it is a variable region. This lack of structure allows the region to recruit and bond many different interaction partners. This region also has the capacity to transactivate transcription without binding estrogen.
The DBD binds estrogen response elements (ERE) of target genes and recruits coactivator proteins responsible for the transcription of these genes. The ERE consist of a palindromic inverted repeat 5'GGTCAnnnTGACC-3' of target genes. The DBD is a highly conserved region. It is composed of 2 C4-type Zn fingers each containing 4 Cys residues coordinating to a Zn atom.
The hinge region connects the DBD and LBD.
LBD binds estrogen, coregulatory proteins, corepressors and coactivators. Genistein is not generated by the endocrine system that binds ERβ like estrogen; both ligands are completely buried within the hydrophobic core
(Hydrophobic, Polar) of the ERβ complex.
Binding at the LBD activates transcription mediated by the DBD. This domain is entirely helical; the LBD interacts with genistein through helices. The conformationally dynamic portion of this region gives rise to ERβ’s ligand-dependent transcriptional activation (AF-2) function. A key element of AF-2 is helix 12 (H12), which acts as a conformational switch; different receptor ligands influence the orientation of H12. Agonist ligands like genistein position H12 across the ligand-binding pocket of the LBD, which provides a coactivator docking surface. Geinstein binding allows the helices of AF-2 to form a shallow hydrophobic binding site for leucine-rich motifs of coactivators to bind. This conformation provides optimal interaction with coactivators and transcription is activated.
Genistein's bicyclic form allows it to hydrogen bond on opposite sides with the hydroxyls of the histidine groups on the receptor. His475's binding to the receptor causes a conformational change and activates the receptor resulting in up-regulation for coactivators. Down-regulation will occur in the presence of corepressor as they bind to repressors and indirectly regulate gene expression. In order for the estrogen receptor β genistein to bind to a receptor and activate it there must be stabilization by a coactivator. The coactivator increases the gene expression and with this increase allows it to bind to an activator group consisting of a DNA binding domain. The estrogen receptor is found to be comprised of a dimer attached to a ligand and coactivator peptide which helps to stabilize the structure of each monomer. The conformational state of helix-12 can be modified by the binding of the coactivator.
This scene depicts the hydrophobic and hydrophilic residues of the estrogen receptor. The hydrophobic regions are primarily on the inside of the protein surrounding genistein (red). Having the hydrophobic residues surrounding the binding pocket will stabilize the structure. The structure of this pocket is tertiary and do to the hydrophobic interactions inside the pocket and hydrophilic interactions on the outside help to stabilize this tertiary structure. The binding pocket is hydrophobic which means that an increase in lipophilicity would increase the affinity for ligands which in this case is genistein. The genistein structure has 3 hydroxyl groups, an ether and an ester. These 3 functional groups are polar and have many possibilities for hydrogen bonding. The His475 and Met336 residues in the binding pocket are capable of forming hydrogen bonds with genistein do to the many hydrogen bond forming functional groups. These residues are different from the residues found in ERα and so the selectivity of genistein is much greater for ERβ.
Upon visualizing the estrogen receptor in an arrow representation, the structure can be classified as parallel or anti-parallel. Here is the zoomed primarily hydrophobic pocket.
Binding of nuclear receptor corepressor 2 peptide and 4-hydroxytamoxifen to human estrogen-related receptor γ. The chemotherapeutic drugs bisphenol and tamoxifen are nestled between 4 alpha helices in the ERR active site.
Estrone
Substrates, such as estrone sulfate, form hydrogen bonds and stacking interactions with residues from each subunit in Cavity 1 of ABCG2 multidrug transporter.
Estradiol
Estriol
Estetrol
- 3l03 - Crystal Structure of human Estrogen Receptor alpha Ligand-Binding Domain in complex with a Glucocorticoid Receptor Interacting Protein 1 Nr Box II peptide and Estetrol (Estra-1,3,5(10)-triene-3,15 alpha,16alpha,17beta-tetrol)
- ↑ Li MJ, Greenblatt HM, Dym O, Albeck S, Pais A, Gunanathan C, Milstein D, Degani H, Sussman JL. Structure of estradiol metal chelate and estrogen receptor complex: The basis for designing a new class of selective estrogen receptor modulators. J Med Chem. 2011 Apr 7. PMID:21473635 doi:10.1021/jm200192y
