Human antigen R (HuR) functions as a major post-transcriptional regulator of gene expression through its RNA-binding activity. HuR is composed by three RNA recognition motifs, namely RRM1, RRM2, and RRM3. The two N-terminal RRM domains are disposed in tandem and contribute mostly to HuR interaction with adenine and uracil-rich elements (ARE) in mRNA. Here, we used a combination of NMR and electrospray ionizationā??ion mobility spectrometryā??mass spectrometry (ESIā??IMSā??MS) to characterize the structure, dynamics, RNA recognition, and dimerization of HuR RRM1. Our solution structure reveals a canonical RRM fold containing a 19-residue, intrinsically disordered N-terminal extension, which is not involved in RNA binding. NMR titration results confirm the primary RNA-binding site to the two central Ī²-strands, Ī²1 and Ī²3, for a cyclooxygenase 2 (Cox2) ARE I-derived, 7-nucleotide RNA ligand. We show by 15N relaxation that, in addition to the N- and C-termini, the Ī²2ā??Ī²3 loop undergoes fast backbone dynamics (psā??ns) both in the free and RNA-bound state, indicating that no structural ordering happens upon RNA interaction. ESIā??IMSā??MS reveals that HuR RRM1 dimerizes, however dimer population represents a minority. Dimerization occurs via the Ī±-helical surface, which is oppositely orientated to the RNA-binding Ī²-sheet. By using a DNA analog of the Cox2 ARE I, we show that DNA binding stabilizes HuR RRM1 monomer and shifts the monomerā??dimer equilibrium toward the monomeric species. Altogether, our results deepen the current understanding of the mechanism of RNA recognition employed by HuR.
[NMR paper] Solution NMR Studies of the Ligand-Binding Domain of an Orphan Nuclear Receptor Reveals a Dynamic Helix in the Ligand-Binding Pocket.
Solution NMR Studies of the Ligand-Binding Domain of an Orphan Nuclear Receptor Reveals a Dynamic Helix in the Ligand-Binding Pocket.
Solution NMR Studies of the Ligand-Binding Domain of an Orphan Nuclear Receptor Reveals a Dynamic Helix in the Ligand-Binding Pocket.
Biochemistry. 2018 Mar 16;:
Authors: Daffern N, Chen Z, Zhang Y, Pick L, Radhakrishnan I
Abstract
The ligand-binding domains (LBD) of the NR5A subfamily of nuclear receptors activate transcription via ligand-dependent and ligand-independent mechanisms. The...
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[NMR paper] Systematic solution to homo-oligomeric structures determined by NMR.
Systematic solution to homo-oligomeric structures determined by NMR.
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Protein structure determination by NMR has predominantly relied on simulated annealing-based conformational search for a converged fold using primarily...
[NMR paper] NMR study of the Z-DNA binding mode and B-Z transition activity of the Z? domain of human ADAR1 when perturbed by mutation on the ?3 helix and ?-hairpin.
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Arch Biochem Biophys. 2014 Jul 7;
Authors: Jeong M, Lee AR, Kim HE, Choi YG, Choi BS, Lee JH
Abstract
The Z? domains of human ADAR1 (Z?ADAR1) bind to Z-DNA via interaction mediated by the ?3-core and ?-hairpin....
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[NMR paper] Unique Structure and Dynamics of the EphA5 Ligand Binding Domain Mediate Its Binding Specificity as Revealed by X-ray Crystallography, NMR and MD Simulations.
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PLoS One. 2013;8(9):e74040
Authors: Huan X, Shi J, Lim L, Mitra S, Zhu W, Qin H, Pasquale EB, Song J
Abstract
The 16 EphA and EphB receptors represent the largest family of receptor tyrosine kinases, and their interactions...
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[NMR paper] Solution Structure, Dynamics and Binding Studies of a Family 11 Carbohydrate-Binding Module from Clostridium thermocellum (CtCBM11).
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Biochem J. 2013 Jan 29;
Authors: Viegas A, Sardinha J, Freire F, Duarte DF, Carvalho AL, Fontes CM, Romćo MJ, Macedo AL, Cabrita EJ
Abstract
Non-catalytic cellulosomal carbohydrate-binding modules (CBMs) are responsible for increasing the catalytic efficiency of...
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[NMR paper] NMR investigation on the DNA binding and B-Z transition pathway of the Z? domain of human ADAR1.
NMR investigation on the DNA binding and B-Z transition pathway of the Z? domain of human ADAR1.
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Abstract
Human ADAR1, which has two left-handed Z-DNA binding domains, preferentially binds Z-DNA rather than B-DNA with a high...
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[NMR paper] NMR solution structure of a peptide from the mdm-2 binding domain of the p53 protein
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We have recently found that a peptide from the mdm-2 binding domain of the p53 protein induced rapid membranolytic necrosis of a variety of different human...
Oligomeric transition and dynamics of RNA binding by the HuR RRM1 domain in solution
Linear Mode
