Estrogen Receptor Folding Modulates cSrc Kinase SH2 Interaction via a Helical Binding Mode.
The estrogen receptors (ERs) feature, next to their transcriptional role, important nongenomic signaling actions, with emerging clinical relevance. The Src Homology 2 (SH2) domain mediated interaction between cSrc kinase and ER plays a key role in this; however the molecular determinants of this interaction have not been elucidated. Here, we used phosphorylated ER peptide and semisynthetic protein constructs in a combined biochemical and structural study to, for the first time, provide a quantitative and structural characterization of the cSrc SH2-ER interaction. Fluorescence polarization experiments delineated the SH2 binding motif in the ER sequence. Chemical shift perturbation analysis by nuclear magnetic resonance (NMR) together with molecular dynamics (MD) simulations allowed us to put forward a 3D model of the ER-SH2 interaction. The structural basis of this protein-protein interaction has been compared with that of the high affinity SH2 binding sequence GpYEEI. The ER features a different binding mode from that of the "two-pronged plug two-hole socket" model in the so-called specificity determining region. This alternative binding mode is modulated via the folding of ER helix 12, a structural element directly C-terminal of the key phosphorylated tyrosine. The present findings provide novel molecular entries for understanding nongenomic ER signaling and targeting the corresponding disease states.
Laboratory of Chemical Biology, Department of Biomedical Engineering and Institute of Complex Molecular Systems, Eindhoven University of Technology , 5612AZ Eindhoven, The Netherlands.
Lidia
L
Nieto
Laboratory of Chemical Biology, Department of Biomedical Engineering and Institute of Complex Molecular Systems, Eindhoven University of Technology , 5612AZ Eindhoven, The Netherlands.
Inga M
IM
Tharun
Laboratory of Chemical Biology, Department of Biomedical Engineering and Institute of Complex Molecular Systems, Eindhoven University of Technology , 5612AZ Eindhoven, The Netherlands.
Mark
M
Balk
Bijvoet Center for Biomolecular Research, NMR Spectroscopy Utrecht University , 3584CH Utrecht, The Netherlands.
Hans
H
Wienk
Bijvoet Center for Biomolecular Research, NMR Spectroscopy Utrecht University , 3584CH Utrecht, The Netherlands.
Rolf
R
Boelens
Laboratory of Chemical Biology, Department of Biomedical Engineering and Institute of Complex Molecular Systems, Eindhoven University of Technology , 5612AZ Eindhoven, The Netherlands.
Christian
C
Ottmann
Laboratory of Chemical Biology, Department of Biomedical Engineering and Institute of Complex Molecular Systems, Eindhoven University of Technology , 5612AZ Eindhoven, The Netherlands.
Lech-Gustav
LG
Milroy
Laboratory of Chemical Biology, Department of Biomedical Engineering and Institute of Complex Molecular Systems, Eindhoven University of Technology , 5612AZ Eindhoven, The Netherlands.
Luc
L
Brunsveld
D000595:Amino Acid Sequence; D001665:Binding Sites; D005260:Female; D005453:Fluorescence; D006801:Humans; D009682:Magnetic Resonance Spectroscopy; D008954:Models, Biological; D056004:Molecular Dynamics Simulation; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry / Q000235:genetics; D011485:Protein Binding; D017510:Protein Folding; D011960:Receptors, Estrogen / Q000737:chemistry / Q000378:metabolism*; D018909:src Homology Domains; D019061:src-Family Kinases / Q000737:chemistry / Q000378:metabolism*
D010455:Peptides; D011960:Receptors, Estrogen; D019061:src-Family Kinases