Homo- and heteronuclear NMR studies of the human retinoic acid receptor beta DNA-binding domain: sequential assignments and identification of secondary structure elements.
An 80 amino acid polypeptide corresponding to the DNA-binding domain (DBD) of the human retinoic acid receptor beta (hRAR-beta) has been studied by 1H homonuclear and 15N-1H heteronuclear two- and three-dimensional (2D and 3D) NMR spectroscopy. The polypeptide has two putative zinc fingers homologous to those of the receptors for steroid and thyroid hormones and vitamin D3. The backbone 1H resonances as well as over 90% of the side-chain 1H resonances have been assigned by 1H homonuclear 2D techniques except for the three N-terminal residues. The assignments have been confirmed further by means of 15N-1H heteronuclear 3D techniques, which also yielded the assignments of the 15N resonances. Additionally, stereospecific assignments of methyl groups of five valine residues were made. Sequential and medium-range NOE connectivities indicate several elements of secondary structure including two alpha-helices consisting of residues E26-Q37 and Q61-E70, a short antiparallel beta-sheet consisting of residues P7-F9 and S23-C25, four turns consisting of residues P7-V10, I36-N39, D47-C50, and F69-G72, and several regions of extended peptide conformation. Similarly, two helices are found in the glucocorticoid receptor (GR) DBD in solution [Härd et al. (1990) Science 249, 157-160] and in crystal [Luisi et al. (1991) Nature 352, 497-505], and in the estrogen receptor (ER) DBD in solution [Schwabe et al. (1990) Nature 348, 458-461], although the exact positions and sizes of the helices differ somewhat. Furthermore, long-range NOEs suggest the existence of a hydrophobic core formed by the two helices.
Katahira
M
M
Bijvoet Center for Biomolecular Research, University of Utrecht, The Netherlands.
Knegtel
R M
RM
Boelens
R
R
Eib
D
D
Schilthuis
J G
JG
van der Saag
P T
PT
Kaptein
R
R
D000595:Amino Acid Sequence; D002352:Carrier Proteins / Q000737:chemistry*; D004268:DNA-Binding Proteins / Q000737:chemistry*; D006801:Humans; D006860:Hydrogen Bonding; D006863:Hydrogen-Ion Concentration; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry; D011487:Protein Conformation; D011960:Receptors, Estrogen / Q000737:chemistry; D011965:Receptors, Glucocorticoid / Q000737:chemistry; D018168:Receptors, Retinoic Acid; D016415:Sequence Alignment; D013696:Temperature; D035582:Transcription Factor TFIIIA; D014157:Transcription Factors / Q000737:chemistry; D014212:Tretinoin / Q000378:metabolism; D016335:Zinc Fingers
D002352:Carrier Proteins; D004268:DNA-Binding Proteins; D010446:Peptide Fragments; D011960:Receptors, Estrogen; D011965:Receptors, Glucocorticoid; D018168:Receptors, Retinoic Acid; D035582:Transcription Factor TFIIIA; D014157:Transcription Factors; D014212:Tretinoin