Mid-IR quantum cascade laser spectroscopy to resolve lipid dynamics during the photocycle of bacteriorhodopsin

Membranes are crucial for the functionality of membrane proteins in several cellular processes. Time-resolved infrared (IR) spectroscopy enables the investigation of interaction-induced dynamics of the protein and the lipid membrane. The photoreceptor and proton pump bacteriorhodopsin (BR) was reconstituted into liposomes, mimicking the native purple membrane. By utilization of deuterated lipid alkyl chains, corresponding vibrational modes are frequency-shifted into a spectrally silent window that allows us to monitor lipid dynamics during the photoreaction of BR. Our home-built quantum cascade laser (QCL)-based IR spectrometer covers all relevant spectral regions to detect both lipid and protein vibrational modes. QCL-probed transients at single wavenumbers are compared with the previously performed step-scan Fourier-transform IR measurements. The absorbance changes of the lipids could be resolved by QCL-measurements with a much better signal-to-noise ratio and with nanosecond time resolution. We found a correlation of the lipid dynamics with the protonation dynamics in the M intermediate. QCL spectroscopy extends the study of the protein's photocycle toward dynamics of the interacting membrane.

Automated summary

Membranes are crucial for the functionality of membrane proteins, and time-resolved infrared spectroscopy enables the investigation of interaction-induced dynamics of the protein and the lipid membrane. By utilizing deuterated lipid alkyl chains, we can monitor lipid dynamics during the photoreaction of bacteriorhodopsin. Our home-built quantum cascade laser-based IR spectrometer covers all relevant spectral regions and allows for better signal-to-noise ratio and nanosecond time resolution. QCL spectroscopy extends the study of the protein's photocycle toward dynamics of the interacting membrane.