title: Structure refinement of the glucocorticoid receptor-DNA binding domain from NMR data by relaxation matrix calculations.
abstract: The solution structure of the glucocorticoid receptor (GR) DNA-binding domain (DBD), consisting of 93 residues, has been refined from two and three-dimensional NMR data using an ensemble iterative relaxation matrix approach followed by direct NOE refinement with DINOSAUR. A set of 47 structures of the rat GR fragment Cys440-Arg510 was generated with distance geometry and further refined with a combination of restrained energy minimization and restrained molecular dynamics in a parallel refinement protocol. Distance constraints were obtained from an extensive set of NOE build-up curves in H2O and 2H2O via relaxation matrix calculations (1186 distance constraints from NOE intensities, 10 phi and 22 chi 1 dihedral angle constraints from J- coupling data were used for the calculations). The root-mean-square deviation values of the 11 best structures on the well-determined part of the protein (Cys440 to Ser448, His451 to Glu469 and Pro493 to Glu508) are 0.60 A and 1.20 A from the average for backbone and all heavy atoms, respectively. The final structures have R-factors around 0.40 and good stereochemical qualities. The first zinc-coordinating domain of the GR DBD is very similar to the crystal structure with a root-mean-square difference of 1.4 A. The second zinc-coordinating domain is still disordered in solution. No secondary structure element is found in this domain in the free state. As suggested by crystallographic studies on the estrogen receptor DBD-DNA and GR DBD-DNA complexes, part of this region will form a distorted helix and the D-box will undergo a conformational change upon cooperative binding to DNA.
authors.affiliation: Bijvoet Center for Biomolecular Research Utrecht University, The Netherlands.
authors.forename: M A
authors.initials: MA
authors.lastname: van Tilborg
authors.forename: A M
authors.initials: AM
authors.lastname: Bonvin
authors.forename: K
authors.initials: K
authors.lastname: Hård
authors.forename: A L
authors.initials: AL
authors.lastname: Davis
authors.forename: B
authors.initials: B
authors.lastname: Maler
authors.forename: R
authors.initials: R
authors.lastname: Boelens
authors.forename: K R
authors.initials: KR
authors.lastname: Yamamoto
authors.forename: R
authors.initials: R
authors.lastname: Kaptein
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D004268:DNA-Binding Proteins / Q000737:chemistry*; D009682:Magnetic Resonance Spectroscopy; D008961:Models, Structural; D008969:Molecular Sequence Data; D051381:Rats; D011965:Receptors, Glucocorticoid / Q000737:chemistry*; D016335:Zinc Fingers
chemical_list: D004268:DNA-Binding Proteins; D011965:Receptors, Glucocorticoid