Dr. Mengyang Xu

During the last century, science and technology based on terahertz (THz) frequency electromagnetic radiation have developed rapidly, to the extent that it now gives valuable insight in the fields of biophysics, medical applications, materials science, spaceborne technologies, security imaging systems and telecommunication. In this dissertation, I present the investigation of protein intramolecular long- range vibrations through optical studies of various proteins using THz time domain spectroscopy (THz TDS), Anisotropic Terahertz Microscopy (ATM), and Polarization-varying Anisotropic Terahertz Microscopy (PV-ATM). The biggest challenge to characterize protein dynamics is the overlap of large water absorption and protein vibrational response, which is overcome by the decomposition of the THz absorbance spectrum. The understanding of structural dynamics in biochemistry by THz measurements is still in its infancy. Here I will discuss spectroscopic methods and computational techniques that were developed for extracting protein dynamics from the solvent dynamics. The THz absorbance can be decomposed into protein intramolecular vibrations and solvent intermolecular vibrations through resonance fitting for the low temperature THz time domain spectroscopy (THz TDS) measurements. The systematic study on the protein structural dynamics and its correlation with thermal stability, and photobleaching for red fluorescent proteins (RFPs) was completed by the temperature dependent THz TDS. We find a clear correlation between picosecond flexibility and thermal stability suggesting THz TDS as a quick nondestructive measure of structural stability. We also find that the protein dynamical transition is shifted up from the solvent dynamical turn on. The shift in protein dynamical turn-on temperature relative to the solvent suggests an increase in the collectivity of motions which requires additional mobile waters in the photobleached state. This enhanced internal coupling is consistent with the observed increase in thermal stability. ATM technique can isolate the protein vibrations with specific orientations from the solvent isotropic background by the anisotropic measurements. Instead of altering the orientation of the naturally aligned protein crystals, another way to extract protein dynamics from the water is to continuously change THz polarization, which introduces a modified but much less time-consuming technique, PV-ATM. The temperature dependence in PV-ATM results on the benchmarking protein, chicken egg white lysozyme (CEWL), shows how protein motions escape the water cage due to the solvent immobility at low temperatures. Moreover, the temperature dependent PV-ATM measurements on Fenna-Matthews-Olson protein (FMO) demonstrate that the protein long-range vibrations have a significant effect on the excitation energy transfer and excess energy dissipation in photosynthetic systems.

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