Use this URL to cite or link to this record in EThOS: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.675253
Title: Accessing ultrafast protein dynamics through 2DIR spectroscopy of intrinsic ligand vibrations
Author: Simpson, Niall
ISNI:       0000 0004 5370 8651
Awarding Body: University of Strathclyde
Current Institution: University of Strathclyde
Date of Award: 2015
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Abstract:
Proteins are complex molecular machines that facilitate the chemical reactions fundamental to life. Their functions are encoded in a linear sequence of amino acids, of which only 20 species are found in nature. Yet the functional and structural diversity accessible through these building blocks is vast. Molecular and atomic-level protein studies have been crucial to our understanding of health and treatment of disease, with increasingly sophisticated experimental and computational methods continuing to provide new information with which to advance medicine. However, the requirement for more detailed understanding of proteins has risen through the emergence of multi-antibiotic-resistant bacteria and also through the potential to design synthetic proteins of novel function. Paradigms of protein function have evolved significantly since early studies, though few all-encompassing descriptions have been proposed, owing to the complex, dynamic structures of these large biomolecules. Presently, the relationship between protein structural motions at different timescales appears to hold vital significance to the elusive aspects of biological mechanisms. No single measurement technique is capable of accessing the multitude of timescales over which protein motions occur, and thus concerted investigation is necessary. Observation of dynamics at the femtosecond-picosecond timescale has only recently become possible through the development of new experimental techniques, allowing a new class of protein motions to be investigated. In this thesis, the advanced technique of two-dimensional infrared spectroscopy (2DIR) is employed to study three biomolecular systems with implications to ubiquitous protein interactions. The aims of these investigations are, firstly, to demonstrate the suitability of 2DIR spectroscopy in gathering novel dynamic information from biological systems that is not accessible via other methods, and secondly, to derive the potential physical significance of these dynamics as they relate to biological function. A description of the underlying theory of 2DIR is presented in this Chapter, along with the considerations that must be made in the application of such a technique to complex biological case-studies. In Chapter (2), descriptions are given for the experimental setups used to acquire infrared spectra, specifically, Fourier transform infrared (FTIR), pump-probe and 2DIR spectroscopies. In Chapter (3) the catalytic-site dynamics of two closely-related haem proteins are each studied by monitoring the vibrational evolution of a nitric oxide (NO) probe molecule bound to the haem centre. A comparison of the active site dynamics is performed in order to correlate the observed differences with discrepancies between the protein reaction mechanisms. Chapter (4) explores the potential of a coenzyme with high protein-binding promiscuity to serve as an intrinsic reporter of the dynamics that occur at substrate binding sites. Infrared analysis and categorisation of the free coenzyme molecule is performed in order to establish its effectiveness as a probe. In Chapter (5), method-development strategies are proposed for the extraction of 2DIR data from large, complex protein-protein systems, with the objective of expanding the range of interactions on which 2DIR can effectively report. Both well-established and novel strategies are employed, and the potential and limitations of the technique are discussed in the context of these demanding case-studies. Chapter (6) draws together conclusions and an overview of progress made and discusses future directions.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.675253  DOI: Not available
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