Secondary structure changes of Bax at detergent concentration above CMC_Brij-35, probed by far-UV CD.

<p>Spectra of 5 µM Bax in the absence (–) and presence (–) of 0.05% Brij-35, in 25 mM sodium phosphate (pH 7.4), 50 mM NaCl, 1 mM EDTA, and 1 mM DTT at 20°C.</p

Far-UV CD spectra of Bcl-2 and Bax, and thermal unfolding of the latter, at detergent concentration below CMC_Brij-35.

<p>(<b>A</b>) Spectra of 5 µM Bax in the absence (–) and presence (–) of 0.0028% Brij-35, in 25 mM sodium phosphate (pH 7.4), 50 mM NaCl, 1 mM EDTA, and 1 mM DTT, at 20°C. (<b>B</b>) Thermal unfolding of 5 µM Bax in the absence (–) and presence (–) of 0.0028% Brij-35. (<b>C</b>) Far-UV CD spectra of 5 µM Bcl-2 in 0.0028% (–) and 0.05% (–) Brij-35, at 20°C. The composition of the buffer used in experiments (<b>B</b>) and (<b>C</b>) was the same as for (<b>A</b>).</p

Studying the interplay between Bax and Bcl-2 at detergent concentration above CMC_Brij-35.

<p>(<b>A</b>) Far-UV CD spectra of a solution containing 5 µM Bax+5 µM Bcl-2 (experimental, (–)) and the sum of their individual spectra (theoretical, (–)), in the presence of 25 mM sodium phosphate (pH 7.4), 50 mM NaCl, 1 mM EDTA, 1 mM DTT and 0.05% Brij-35, at 20°C. (<b>B</b>) Thermal unfolding of 5 µM Bax+5 µM Bcl-2 (experimental, (–)) compared to the sum of the melting diagrams for the individual proteins (theoretical, (–)). (<b>C</b>) Normalized fluorescence spectra of 2.5 µM antimycin A₂ alone (··) and when incubated with 2.5 µM Bax (­­), 2.5 µM Bcl-2 (–) or both Bax and Bcl-2 (–), at 23°C. The composition of the buffer used in experiments (<b>B</b>) and (<b>C</b>) was the same as for (<b>A</b>).</p

Studying the interplay between Bax and Bcl-2 at detergent concentration below CMC_Brij-35.

<p>(<b>A</b>) Far-UV CD spectra of 5 µM Bax+5 µM Bcl-2 (experimental, (–)) and the sum of their individual spectra (theoretical, (–)), in the presence of 25 mM sodium phosphate (pH 7.4), 50 mM NaCl, 1 mM EDTA, 1 mM DTT, and 0.0028% Brij-35, at 20°C. (<b>B</b>) Thermal unfolding of 5 µM Bax+5 µM Bcl-2 (experimental, (–)), and summed diagrams for the melting of the individual proteins (theoretical, (–)). (<b>C</b>) Normalized fluorescence spectra of 2.5 µM antimycin A₂ alone (··) and when incubated together with 2.5 µM Bax (­­), 2.5 µM Bcl-2 (–) or both Bax and Bcl-2 (–), at 23°C. The composition of the buffer used in experiments (<b>B</b>) and (<b>C</b>) was the same as for (<b>A</b>).</p

Zoomed region of the ¹⁵N-HSQC of CD79a in different conditions.

<p>(<b>A</b>) NaPi buffer, (<b>B</b>) 6 M urea, (<b>C</b>) 20% TFE, (<b>D</b>) reduced spin label (MTSL) attached to CD79a, (<b>E</b>) K4C/C35S form, (<b>F</b>) Y25E/Y36E form. Selected peaks are annotated to show rearrangements of the signal position for the different conditions (19Y, 35M) or position of the specific mutations (4K, 33C, 25Y, 36Y). Peaks outside the zoomed region are shown as arrows pointing towards the correct position.</p

Cell-free expressed cytosolic constructs of the T cell- and B cell receptor.

<p>(<b>A</b>) Cartoon of the receptors with indicated immunoreceptor tyrosine-based activation motifs (ITAMs), (<b>B</b>) SDS-PAGE gel of <i>in vitro</i> expressed disordered constructs in levels suitable for NMR experiments.</p

Assignment validation procedure with jackknife resampling.

<p>See text for explanations.</p

Progress of targeted acquisition versus total measurement time for a 120 µM sample of CD79a.

<p>Build-ups are shown for the number of assigned residues and the number of detected peaks in individual BEST-TROSY-type experiments <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0062947#pone.0062947-Favier1" target="_blank">[14]</a>.</p