Volume 113, Issue 6, Pages 1290-1300 (September 2017) pH-Dependent Membrane Interactions of the Histidine-Rich Cell-Penetrating Peptide LAH4-L1  Justine Wolf, Christopher Aisenbrey, Nicole Harmouche, Jesus Raya, Philippe Bertani, Natalia Voievoda, Regine Süss, Burkhard Bechinger  Biophysical Journal  Volume 113, Issue 6, Pages 1290-1300 (September 2017) DOI: 10.1016/j.bpj.2017.06.053 Copyright © 2017 Biophysical Society Terms and Conditions

Figure 1 31P solid-state NMR spectra of POPC/POPS 3:1 membranes acquired at 11.8 Tesla in the absence (A and F) and presence of increasing amounts of LAH4-L1 at pH 7.4 (A–E) and pH 5 (F–K). The P/L ratios are indicated. The temperature was set to 30°C (303 K) except for the spectrum in (I), which was recorded at 16°C. At 30°C, this same sample exhibits an isotropic resonance (data not shown). Biophysical Journal 2017 113, 1290-1300DOI: (10.1016/j.bpj.2017.06.053) Copyright © 2017 Biophysical Society Terms and Conditions

Figure 2 31P solid-state NMR spectra of DMPC membranes acquired at 11.8 Tesla in the absence (A and H) and presence of increasing amounts of LAH4-L1 at pH 7.4 (A–G) and pH 5 (H–O). The P/L ratios are indicated. The temperature was set to 37°C (310 K; first and third columns) or 7°C (280 K; second and fourth columns). Biophysical Journal 2017 113, 1290-1300DOI: (10.1016/j.bpj.2017.06.053) Copyright © 2017 Biophysical Society Terms and Conditions

Figure 3 The δ⊥ value of POPC/POPS 3:1 and DMPC membranes as a function of LAH4-L1/lipid molar ratio extracted from the 31P solid-state NMR spectra shown in Figs. 1 and 2. The temperature was set to 30°C (303 K) for POPC/POPS and to 37°C (310 K) for DMPC. Biophysical Journal 2017 113, 1290-1300DOI: (10.1016/j.bpj.2017.06.053) Copyright © 2017 Biophysical Society Terms and Conditions

Figure 4 2H solid-state NMR spectra acquired at 11.8 Tesla for POPC/2H31-POPS 3:1 in the absence (A and F) and presence of increasing amounts of LAH4-L1 at pH 7.4 (A–E) and pH 5 (F–K). The P/L ratios are indicated. The temperature was set to 30°C (303 K), except for (I), where it was set to 16°C (289 K). Biophysical Journal 2017 113, 1290-1300DOI: (10.1016/j.bpj.2017.06.053) Copyright © 2017 Biophysical Society Terms and Conditions

Figure 5 The deuterium order parameter as a function of carbon position of POPC/2H31-POPS 3:1 (A and B) and 2H54-DMPC (C and D) in the presence of increasing amounts of LAH4-L1 at pH 7.4 (A and C) or at pH 5 (B and D). The temperature was set to 30°C (303 K) for POPC/POPS and 37°C (310 K) for DMPC. To see this figure in color, go online. Biophysical Journal 2017 113, 1290-1300DOI: (10.1016/j.bpj.2017.06.053) Copyright © 2017 Biophysical Society Terms and Conditions

Figure 6 Proton-decoupled 15N (A–D) and corresponding 31P solid-state NMR spectra (E–H) of [15N-Leu14]-LAH4-L1 reconstituted in POPC/POPS 3:1 mol/mole bilayers oriented with the membrane normal parallel to the magnetic field direction. Spectra in the first and third columns are at pH 7.4 (A, B, E, and F) and those in the second and last column are at pH 5 (C, D, G, and H). The P/L ratio is 1:50 in the first row, (A, C, E, and G) and 1:25 in the second row (B, D, F, and H). Biophysical Journal 2017 113, 1290-1300DOI: (10.1016/j.bpj.2017.06.053) Copyright © 2017 Biophysical Society Terms and Conditions