Sub-terahertz circular dichroism spectroscopy of solvated proteins via vector network analysis and time-domain spectroscopy

The study of solvated proteins by terahertz (THz) frequency electromagnetic radiation, characterized by submillimeter wavelength signal beams, suffers from strong absorption by water. Yet, water is vital to normal protein function; hydration plays a formative role in a protein acquiring its operational molecular structure. To decouple the traditional conflation of water spectra with that contributed by the protein in conventional transmission or absorption spectroscopy, this thesis innovates the experimental methodology of circular dichroism (CD) spectroscopy; water being achiral. Dealing with the heavy signal loss from absorption by water is countered by the gain afforded by using a state-of-the-art vector network analyser (VNA), that is interfaced with a quasi-optical (QO) circuit, containing the sample of dissolved protein. This apparatus has been employed to co-study CD protein conformational changes conventionally acquired in the far-UV at the discrete frequency of 222 nm. This thesis explores the first acquisition of a continuous THz CD spectra of a protein as it responds to variation in the temperature, salinity and pH of its host aqueous medium, in order to lay foundations for interpreting associated spectra in relation to the given conformational state the protein adopts in response. Experimental work in this thesis was performed over the mm and sub-mm waveguide bands: WR7 (110 - 170 GHz) and WR3 (220 - 325 GHz), respectively; and, THz Time-Domain Spectroscopy (TDS). (200 - 1400 GHz). A QO-based, sub-THz CD spectrometer is demonstrated on the sample protein, myoglobin, over the waveguide bands of WR7 and WR3. In so doing, an equivalent methodology is propounded for acquiring cross-polar spectra that eliminates the need for having to physically rotate the receive-horn through 90˚. A significant source of systematic-error is thereby eliminated (and, no less, random-errors, that may occur when cabling is moved). The 'zero-crossing' points of CD spectra are characterized as being correlated with traditional far-UV CD spectra in the given environmental parameter settings of, for example, pH and/or sample concentration level. For the extending application, the sub-THz CD QO system studies three solvated proteins namely, Myoglobin, Bovine Serum Albumins, and Papain at WR3. Water is used as a buffer, since it contributes no CD signal. Secondary structures of the proteins consist of different amounts of α-helices and β-sheet forms. Different optical path-lengths and concentration levels are considered in order to compare their efficiency in providing sub-THz CD signatures by different structures of proteins. The application of THz TDS is to study the dielectric properties of the three proteins. Dielectric properties of solvated proteins are strongly influenced by the flexibility of protein secondary structures, hydration bonds, and water-protein interaction. By analyzing the protein's dielectric behaviour, explanation of protein CD behaviour will be sought for the sub-THz frequency domain.