External cavity quantum cascade laser vibrational circular dichroism spectroscopy (EC-QCL-VCD) for analysis of proteins at low concentrations

Abstract

Proteins are characterized by their complex three-dimensional structures, which in turn define their function. Characterization of these structures in solution and in a non-destructive way is crucial for being able to gain fundamental understanding of biological processes. One of the possible analytical methods capable of providing this sort of information is vibrational circular dichroism (VCD), which expands the structural sensitivity of classical IR absorbance spectroscopy by the chiral sensitivity of infrared (IR) circular dichroism. While this technique is powerful, it is plagued by low signal intensities and long measurement times. Here we present an optical setup leveraging the high brilliance of a quantum cascade laser to measure proteins in D 2 O at extended pathlengths ( >200 μm). The developed approach uses a dual beam set-up with a balanced detector where in one channel a photoelastic modulator is placed to generate left and right hand circular polarized light. The difference in absorption of the circular polarized light is calculated and displayed as the VCD spectrum of the sample. The obtained results will be compared to established Fourier-transform infrared spectroscopy (FT-IR) in terms of noise levels and in its applicability to secondary structure elucidation of proteins. Protein concentrations as low as 2 mg/mL were accessible by the laser-based system at a measurement time of 1 h