Structure of the carboxy-terminal region of thyroid hormone nuclear receptors and its possible role in hormone-dependent intermolecular interactions.
The thyroid hormone nuclear receptors (TRs) are ligand-dependent transcription factors. To understand the molecular basis of ligand-dependent transactivation, we studied the structure of their carboxy-terminal activation domain. We analyzed the structures of the peptides derived from the C-terminal sequences of human TR subtypes beta 1 (h-TR beta 1) and alpha 1 (h-TR alpha 1) and a human TR mutant, PV, by circular dichroism (CD). Mutant PV has a C-terminal frameshift mutation and does not bind to the thyroid hormone, 3,3',5-triiodo-L-thyronine (T3). Analyses of the secondary structures of the peptides by CD indicate that five amino acids, EVFED, are part of an amphipathic alpha-helix which is required to maintain the structural integrity of the hormone binding domain. A monoclonal antibody, C4 (mAb C4), which recognizes both h-TR beta 1 and h-TR alpha 1 was developed. Using a series of truncated mutants and synthetic peptides, we mapped the epitope of mAb C4 to the conserved C-terminal amino acids, EVFED. Analysis of the binding data indicates that binding of T3 to either h-TR beta 1 or h-TR alpha 1 was competitively inhibited by mAb C4. Deletion of C-terminal amino acids including EVFED led to a total loss of T3 binding activity. Thus, part of the T3 binding site is located in this five amino acid segment. T3 may transduce its hormonal signal to the transcriptional machinery via interaction with EVFED at the C-terminus of TRs.
Bhat
M K
MK
Laboratory of Molecular Biology, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892-4255, USA.
McPhie
P
P
Ting
Y T
YT
Zhu
X G
XG
Cheng
S Y
SY
D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000378:metabolism; D001483:Base Sequence; D001665:Binding Sites; D001667:Binding, Competitive; D015153:Blotting, Western; D002467:Cell Nucleus / Q000737:chemistry*; D002942:Circular Dichroism; D000939:Epitopes; D016368:Frameshift Mutation; D007163:Immunosorbent Techniques; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry* / Q000276:immunology; D017433:Protein Structure, Secondary; D011988:Receptors, Thyroid Hormone / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D014284:Triiodothyronine / Q000378:metabolism / Q000494:pharmacology*
D000911:Antibodies, Monoclonal; D000939:Epitopes; D010446:Peptide Fragments; D011988:Receptors, Thyroid Hormone; D014284:Triiodothyronine