BPA concentration dependance of WaterLOGSY signal.

<p>Signal intensities from the methyl groups of BPA (in green) and the DMSO (in grey) are shown as a function of BPA concentration. The <b>P11</b> peptide concentration is 200 <i>μ</i>M.</p

Domain organization of Androgen Nuclear Receptor.

<p>Schematic representation of AR showing the structured DBD and LBD domains, connected by the hinge region in grey and the N-terminal disordered domain (NTD). A zoom of the <b>P11</b> region shows the sequence of the peptide investigated in this work and the amyloid fibers formed upon Cys240 oxidation.</p

Comparison of BPA WaterLOGSY signal intensities for P11 and polyQ fibers.

<p>The WL signal intensity was measured using the methyl groups of BPA (in green) and DMSO (in grey). The concentration of <b>P11</b> and polyQ peptides was 200 <i>μ</i>M and BPA concentration is 1 mM.</p

WaterLOGSY study of BPA with the P11 amyloid fibers.

<p>(a) WL difference spectrum of a 1 mM solution of BPA in presence of 200 <i>μ</i>M of <b>P11</b> peptide and 10% DMSO-d₆. BPA peaks are labelled, the orange square indicates the methyl resonance of DMSO, and the × indicate two impurities found in the solution, possibly acetonitrile and ter-butanol. b,c) WL difference spectrum of the BPA solution featuring the same composition as in (a) after a first centrifugation step (b) and after a second one (c). Centrifugation was performed at 10000 r.p.m. for 40 min on each occasion. The mixed sign observed on the sharp methyl <b>1</b> signal apparently arises from incomplete signal cancelation during acquisition caused by instrument instabilities.</p

Solution Behavior of the Intrinsically Disordered N‑Terminal Domain of Retinoid X Receptor α in the Context of the Full-Length Protein

Retinoid X receptors (RXRs) are transcription factors with important functions in embryonic development, metabolic processes, differentiation, and apoptosis. A particular feature of RXRs is their ability to act as obligatory heterodimerization partners of class II nuclear receptors. At the same time, these receptors are also able to form homodimers that bind to direct repeat separated by one nucleotide hormone response elements. Since the discovery of RXRs, most of the studies focused on its ligand binding and DNA binding domains, while its N-terminal domain (NTD) harboring a ligand-independent activation function remained poorly characterized. Here, we investigated the solution properties of the NTD of RXRα alone and in the context of the full-length receptor using small-angle X-ray scattering and nuclear magnetic resonance spectroscopy. We report the solution structure of the full-length homodimeric RXRα on DNA and show that the NTD remains highly flexible within this complex

Thermodynamics of Zn²⁺ Binding to Cys₂His₂ and Cys₂HisCys Zinc Fingers and a Cys₄ Transcription Factor Site

The thermodynamics of Zn²⁺ binding to three peptides corresponding to naturally occurring Zn-binding sequences in transcription factors have been quantified with isothermal titration calorimetry (ITC). These peptides, the third zinc finger of Sp1 (Sp1-3), the second zinc finger of myelin transcription factor 1 (MyT1-2), and the second Zn-binding sequence of the DNA-binding domain of glucocorticoid receptor (GR-2), bind Zn²⁺ with Cys₂His₂, Cys₂HisCys, and Cys₄ coordination, respectively. Circular dichroism confirms that Sp1-3 and MyT1-2 have considerable and negligible Zn-stabilized secondary structure, respectively, and indicate only a small amount for GR-2. The p<i>K</i>_a’s of the Sp1-3 cysteines and histidines were determined by NMR and used to estimate the number of protons displaced by Zn²⁺ at pH 7.4. ITC was also used to determine this number, and the two methods agree. Subtraction of buffer contributions to the calorimetric data reveals that all three peptides have a similar affinity for Zn²⁺, which has equal enthalpy and entropy components for Sp1-3 but is more enthalpically disfavored and entropically favored with increasing Cys ligands. The resulting enthalpy–entropy compensation originates from the Zn-Cys coordination, as subtraction of the cysteine deprotonation enthalpy results in a similar Zn²⁺-binding enthalpy for all three peptides, and the binding entropy tracks with the number of displaced protons. Metal and protein components of the binding enthalpy and entropy have been estimated. While dominated by Zn²⁺ coordination to the cysteines and histidines, other residues in the sequence affect the protein contributions that modulate the stability of these motifs