Milanesi, Lilia ORCID: 0000-0002-6369-6610
(2002)
Optical triggers for protein folding.
PhD thesis, University of Sheffield.
Abstract
A better understanding of the mechanism by which protein folding proceeds requires the elucidation of processes that occur on the nanosecond time scale. Advances in laser technology have led to the development of new methods which can initiate and probe protein folding on short time scales. However, as discussed in chapter 1, the current optical methods are applicable to a narrow range of proteins or are limited to the microsecond timescale, which is insufficient to follow many protein rearrangements events.
This thesis describes the development of novel optical triggers that have the potential to be more generally applicable than previous methods for the initiation of protein folding in the submicrosecond time scale.
Chapter 2 outlines two strategies used for the development of our optical trigger mechanisms. Both strategies make use of chemical methods to introduce photocleavable aromatic disulfides at key points of the target protein structure.
Chapter 3 details the synthesis and photolysis of a series of aromatic disulfides that have been used as a model system to test the efficiency of the designed optical trigger mechanism. Our results show that the disulfide bond is cleaved in less than a few picoseconds, thus providing a fast trigger for protein folding.
Chapter 4 details the procedure used to introduce a photolabile aromatic disulfide into the target protein. Preliminary results indicate that the aromatic disulfide crosslink forces the protein in a non-native structure, thus providing a good system for the study of fast folding kinetics.
Chapter 5 details a second procedure that I have used to functionalise the target protein with a photolabile aromatic disulfide attached to a single amino acid residue. Preliminary results indicated that the modification of a single residue does not perturb the protein native structure thus this system is not useful for the study of fast folding kinetics.
Metadata
Supervisors: | Hunter, Christopher Alexander |
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Related URLs: | |
Keywords: | protein-folding, aromatic-disulfides, maleimide, crosslinks, time-resolved spectroscopy |
Awarding institution: | University of Sheffield |
Academic Units: | The University of Sheffield > Faculty of Science (Sheffield) > Chemistry (Sheffield) |
Depositing User: | Dr Lilia Milanesi |
Date Deposited: | 12 Aug 2025 15:30 |
Last Modified: | 12 Aug 2025 15:30 |
Open Archives Initiative ID (OAI ID): | oai:etheses.whiterose.ac.uk:37193 |
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