Modulated orientation sensitive terahertz spectroscopy
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The energies of protein correlated motions lie in the far infrared or THz frequency range (λ = 1 cm – 50 mm, f = 0.03 – 6 THz). The existence of correlated motions has been confirmed by neutron and inelastic x-ray scattering measurements. These techniques require large sample volumes and specialized facilities, limiting their application to systematic studies of changes in correlated motions with functional state and allosteric interactions. Standard terahertz time domain spectroscopy measurements have shown sensitivity to protein-ligand binding, oxidation state, conformation, temperature and hydration. However, the response is broad, in part from the large vibrational density of states and in part from the dielectric response contribution from surface water and side-chains. As an overall strategy to measure the correlated structural motions in protein, we use anisotropic and birefringent behavior of molecular crystals to develop a new technique called MOSTS (Modulated Orientation Sensitive Terahertz Spectroscopy). We achieve high sensitivity and mode separation, by using single molecular crystal such as sucrose and oxalic acid, and rapid modulation of the relative alignment of the terahertz polarization and the crystal axes by rotating the sample. By locking into the signal at the rotation frequency, we determine the polarization sensitive signal and map out the optically active vibrational resonances. To illustrate the technique, we compare our measured spectra with the calculated, and find a close agreement. We measure dielectric properties of oxalic acid, sucrose and protein crystals and polycarbonate sheet using standard terahertz time domain spectroscopy. We determine the absorbances in oxalic acid and sucrose crystals, using MOSTS technique. We compare the resonances in these two distinct methods. Then, we develop a protein model sample by sticking together two thin plates of sucrose and polycarbonate. We carry out standard THz-TDS and MOSTS measurements on the protein model sample. We show that we are able to isolate the vibrational modes from glassy background in protein model sample by using MOSTS.