Protein Conformational and Protonation Dynamics Monitored by Single-Shot IR Spectroscopy

Time-resolved IR spectroscopy is a powerful and label-free technique to study the functional mechanisms of proteins. Its chemical sensitivity allows to follow molecular alterations such as conformational changes or proton transfer steps during the protein reaction. The established method of step-scan FTIR spectroscopy yields spectrally broad information (>1000cm^-1) with sufficiently high time resolution (up to ns) but strictly requires reversible processes and stable samples. Although setups based on tunable quantum cascade lasers (QCL) are capable of collecting kinetic information with ns time resolution by a single acquisition, the spectral information is limited to a single wavelength. To access the whole tuning range (~100cm^-1/Laser) multiple acquisitions are required as well. Contrary, QCL-based dual comb spectroscopy (DCS) allows for a simultaneous broadband (~60cm^-1) observation of reaction kinetics with µs time resolution.

Here, we present time-resolved data in the frequency range of amide I and C=O stretching vibration of carboxylic residues, acquired by single-shot experiments. Our results show that with the use of QCLs it is possible to observe transient conformational changes as well as protonation events of single amino acids within a protein, without any acquisition averaging necessary.