ABSTRACT
We present the structure-based optimization of a series of estrogen receptor-beta (ERbeta) selective ligands. X-ray cocrystal structures of these ligands complexed to both ERalpha and ERbeta are described. We also discuss how molecular modeling was used to take advantage of subtle differences between the two binding cavities in order to optimize selectivity for ERbeta over ERalpha. Quantum chemical calculations are utilized to gain insight into the mechanism of selectivity enhancement. Despite only two relatively conservative residue substitutions in the ligand binding pocket, the most selective compounds have greater than 100-fold selectivity for ERbeta relative to ERalpha when measured using a competitive radioligand binding assay.
Subject(s)
Estrogen Receptor beta/chemistry , Estrogen Receptor beta/metabolism , Amino Acid Sequence , Benzofurans/chemistry , Benzofurans/metabolism , Benzoxazoles/chemistry , Benzoxazoles/metabolism , Binding Sites , Binding, Competitive , Crystallography, X-Ray , Estrogen Receptor alpha/chemistry , Estrogen Receptor alpha/metabolism , Humans , Ligands , Male , Models, Molecular , Molecular Sequence Data , Protein Conformation , Quantum Theory , Radioligand Assay , Structure-Activity Relationship , Substrate SpecificitySubject(s)
GTPase-Activating Proteins/chemistry , Membrane Proteins/chemistry , Nerve Tissue Proteins/chemistry , Nuclear Magnetic Resonance, Biomolecular , Amino Acid Sequence , GTPase-Activating Proteins/genetics , Humans , Internet , Isotopes , Membrane Proteins/genetics , Molecular Sequence Data , Nerve Tissue Proteins/genetics , RGS ProteinsABSTRACT
MAP KAP kinase 2 (MK2), a Ser/Thr kinase, plays a crucial role in the inflammatory process. We have determined the crystal structures of a catalytically active C-terminal deletion form of human MK2, residues 41-364, in complex with staurosporine at 2.7 A and with ADP at 3.2 A, revealing overall structural similarity with other Ser/Thr kinases. Kinetic analysis reveals that the K(m) for ATP is very similar for MK2 41-364 and p38-activated MK2 41-400. Conversely, the catalytic rate and binding for peptide substrate are dramatically reduced in MK2 41-364. However, phosphorylation of MK2 41-364 by p38 restores the V(max) and K(m) for peptide substrate to values comparable to those seen in p38-activated MK2 41-400, suggesting a mechanism for regulation of enzyme activity.
