Golden gate chamber

Attenuated total reflection Fourier-transform infrared spectroscopy (ATR-FTIR) measurements were obtained with a diamond as an internal reflection element (IRE). A drop of lipid solution at 0.5 mg/mL in chloroform was first deposited on the IRE and dried with a nitrogen flux. Then, 5 µL of D₂O from Cambridge Isotope was added on top, and finally, 0.5 µL of Hfq-CTR solution at 20 mg/mL in D₂O was added. The system used was a Bruker Equinox55 purged with dry air and a diamond ATR device with a single reflection at an angle of 45° and closed with a golden gate chamber from Specac (Orpington, UK) to avoid evaporation and non-deuterated water vapor exchange. A spectrum, with a resolution of 4 cm⁻¹, was acquired every 10 min for 15 h. For the polarization study, a gold polarizer was used to select polarization normal to the crystal plane (p-polarization or 0°) and polarization in the crystal plane (s-polarization or 90°). The data were treated with Kinetics, a custom-made program developed in SFMB laboratory (SFMB, Université libre de Bruxelles, Belgique) running under MATLAB (Mathworks, Natick, MA, USA). The water vapor signal was removed by subtracting a reference spectrum of pure water vapor with a coefficient optimized on the amide II area band (1555–1550 cm⁻¹). The spectra were normalized on the ester band from the lipid at 1730 cm⁻¹.

Attenuated total reflection Fourier transform infrared spectroscopy (ATR-FTIR) measurements were obtained with a Bruker Equinox55 spectrometer (Billerica, MA, USA) purged with dry air and a diamond ATR device with a single reflection at an angle of 45° and closed with a golden gate chamber from Specac (Orpington, UK). A total of 5 µL of Hfq-CTR solution at 20 mg/mL in D₂O was deposited. The data were treated with kinetics, a custom-made program (SFMB, Université libre de Bruxelles, Bruxelles, Belgium) running under Matlab 7.5.0 (Mathworks, Natick, MA, USA). The water vapor signal was removed by subtracting a reference spectrum of pure water vapor with a coefficient optimized on the amide II area band (1555–1550 cm⁻¹). To analyze the interaction with lipids, a drop of lipid at 10 mg/mL was deposited on the ATR surface and dried with a nitrogen flux. Then, 5 µL of Hfq-CTR solution at 20 mg/mL was deposited on the lipids. A series of spectra with a resolution of 4 cm⁻¹ were acquired every 10 min overnight.

Attenuated total reflection Fourier-transform infrared spectroscopy (ATR-FTIR) measurements were done on a Bruker Equinox55 spectrophotometer equipped with an MCT detector cooled with liquid nitrogen. 2 µL of the sample was deposited and dried with a nitrogen flow on a Specac diamond and 128 scans were averaged for sample and background. To remove most soluble or small oligomeric Hfq11 peptides (non-polymerized), the deposited sample was washed with a drop of 20 µL of water. The water was removed by pipetting without touching the sample and the sample was finally dried by nitrogen. For measurements of Hfq CTR amyloid structure on lipids, a drop of DOPG lipid solution at 0.5 mg/mL in chloroform (Avanti Polar Lipids, Alabaster, AL, USA) was first deposited on the IRE and dried with a nitrogen flux. 5 µL of D₂O from Cambridge Isotope was added on top and finally, 0.5 µL of assembled Hfq CTR at 20 mg/mL in D₂O was added. The system used was a Bruker Equinox55 purged with dry air and a diamond ATR device with a single reflection at an angle of 45° and closed with a golden gate chamber from Specac (Orpington, UK) to avoid evaporation and non-deuterated water vapor exchange. Spectra with a resolution of 4 cm⁻¹ were acquired every 90 min overnight.