Novel vibrational spectroscopic techniques: long-range order in amyloid fibrils

Amyloid diseases are a major health problem and include neurodegenerative diseases such as Alzheimer's, Parkinson's, and Huntington's disease, as well as type II diabetes and infectious prion diseases, such as Creutzfeldt-Jakob disease (CJD). They are characterised by the deposition of large fibrillar aggregates of protein, both within cells and in the extra cellular matrix, that were thought to cause cell death and result in the observed pathology. The constituent fibres of these fibrils have a common beta-sheet architecture with long-range order despite being able to form from many different proteins. More recently, attention has shifted to precursor soluble oligomer species as the toxic agents in these diseases. However, the precise structure and degree of long-range order in these oligomers has not yet been determined and is difficult to probe using existing biophysical techniques, hampering the development of drugs targeted at this stage of the disease. My Fellowship will make use of terahertz frequency time-domain spectroscopy (THz-TDS) to study long-range order of amyloid fibrils and their precursors. This will be complemented by Raman spectroscopy and molecular modelling calculations to gain an understanding of the inter- and intra-molecular interactions within this important class of biological molecules. THz-TDS is an emerging technique that is proving to be of particular importance for the investigation of organic crystalline compounds. This is because THz spectroscopy has been shown to excite inter-molecular vibrations, thus not only probing chemical composition, but also proving to be extremely sensitive to small changes in crystalline structure. During the Fellowship, I will develop new THz frequency spectroscopic techniques. For example, the inclusion of polarisation sensitive detection will, for the first time at THz frequencies, allow spectroscopic information to be obtained related to the chirality of the molecules, including the secondary structure of proteins and small peptides.

Terahertz (THz) frequency spectroscopy is an emerging technique which has attracted considerable international attention in recent years. My suite of new THz spectroscopy techniques will have wide impact. To give one example, advances in spectral collection, and especially my reflection geometries, will strengthen the use of THz-based techniques for industrial process monitoring and security screening applications. This will be of interest to government agencies including HMGCC, DSTL and HOSDB (with which I have a number of contacts from my PhD research) who have a proven interest in THz imaging and spectroscopy for security applications. No existing THz spectroscopy system (either commercial nor research) can distinguish chiral forms, and so my proposed development of a THz vibrational circular dichroism (VCD) instrument will be of significance to the biological and pharmaceutical communities primarily, along with any industry involved in the use of process analytical techniques, specifically if they are concerned with chiral forms. Direct beneficiaries of this work are clinical end-users, with interest in the development, diagnosis and treatment of amyloid diseases. These are a major health problem and include neurodegenerative diseases such as Alzheimer's, Parkinson's, and Huntington's disease, as well as type II diabetes and infectious prion diseases, such as Creutzfeldt-Jakob disease. They are characterized by deposition of large fibrillar aggregates of protein, although attention is focusing on the precursor soluble oligomer species as the toxic agents. The processes involved in the conversion of these soluble oligomers via nucleation to form amyloid fibrils is poorly understood and is difficult to probe using existing biophysical techniques, hampering drug development. My use of THz spectroscopy to probe the development of long-range order (proven for small molecular systems) of the precursor oligomer species, together with my THz-VCD technique, which could access the underlying secondary/tertiary structure of these systems, providing unique and crucial information in understanding amyloid fibril formation. This combined with an understanding of why so many proteins can form these fibrillar structures, seemingly independently of amino acid sequence, would be a significant step forward in the development of treatments for these debilitating diseases. Leeds has the largest teaching hospital in Europe providing significant access to clinicians as the programme progresses. To disseminate my work widely, results will be published in primary refereed journals with cross-disciplinary readership. The demonstration of THz-VCD of pharmaceutical compounds, and investigations of amyloid fibrils, will be advances publishable in high impact factor journals such as Nature Materials/Biochemistry. Results will also be published at conferences including IRMMW-THz, and ICAVS. I will attend meetings such as those organized by the Royal Society of Chemistry, the Infra-red and Raman Discussion Group, the EPSRC, and Knowledge Transfer Networks (KTNs), to ensure potential beneficiaries find out about and benefit from my research. Finally, IP will be protected by the University Research Support, and the University Knowledge and Innovation Unit, who underpin establishment of research contracts, knowledge-transfer, patents, and licensing issues, together with the University's Venture Capital partner, IP Group.