Functionally relevant solvation dynamics in the protein interior
Abstract/Contents
- Abstract
- Functionally relevant solvation dynamics in the protein interior Stanford University, 2010. The role of solvent in the kinetics and thermodynamics of charge transfer reactions in simple solvents is well studied. However, biologically relevant systems often involve chemical reactions not in simple aqueous environments, but within a protein interior that constitutes an organized solvent environment. The role of the protein environment during catalysis and charge transfer reactions continues to be debated. To determine the role of the protein environment during charge transfer reactions, experiments presented in this dissertation were designed to measure the time-resolved response of a protein environment to sudden electronic perturbation. Specifically, time-resolved fluorescence measurements were used to determine the solvation response of a chromophore bound in the protein interior. An unnatural amino acid containing a fluorescent side chain, aladan, was synthetically incorporated into seven sites of a small globular protein, GB1, in order to determine the degree of solvation in different regions of a single protein. In all regions, the solvation dynamics showed an ultra-fast solvation response on the subpicosecond timescale. However, in addition to the subpicosecond dynamics, buried sites also showed relatively long solvation behavior over nanoseconds. mPlum, a variant of the red-fluorescent protein DsRed, was produced by a directed evolution experiment in the Roger Tsien lab. Ultra-fast fluorescence upconversion experiments measured a solvation response in mPlum. Examining the comprehensive library of mutants generated by the directed evolution experiment identified a single hydrogen bond between glutamic acid 16 and the chromophore to be the origin of the solvation process. The possibility of localizing a solvation response to a single interaction is only possible in an organized solvent system like a protein interior. The catalytic relevance of solvation dynamics was studied with a light-driven reaction analog for isomerisation of [delta]5-3-ketosteroids in ketosteroid isomerase. The light-driven analog consists of a photoacid bound to the active site of ketosteroid isomerase. In the ground state, the photoacid electrostatically resembles the reaction intermediate. In the excited state, the photoacid resembles the starting material. The solvation response recorded from the fluorescing photoacid shows a dramatically reduced solvation capacity and slower dynamics than observed with the same fluorophore in solution. Solvation within the protein occurs on the nanosecond timescale compared to picoseconds in solution. The active site of ketosteroid isomerase does show dynamic electrostatic behavior during the simulated reaction; however, the response is among the slowest recorded. The resistance to electrostatic perturbation suggests an electrostatically preorganized environment which does not significantly reorganize during catalysis.
Description
| Type of resource | text |
|---|---|
| Form | electronic resource; remote; computer; online resource |
| Extent | 1 online resource |
| Publication date | 2010 |
| Issuance | monographic |
| Language | English |
Creators/Contributors
| Associated with | Childs, William James |
|---|---|
| Associated with | Stanford University, Department of Chemistry. |
| Primary advisor | Boxer, Steven G. (Steven George), 1947- |
| Thesis advisor | Boxer, Steven G. (Steven George), 1947- |
| Thesis advisor | Dai, Hongjie, 1966- |
| Thesis advisor | Zare, Richard N |
| Advisor | Dai, Hongjie, 1966- |
| Advisor | Zare, Richard N |
Subjects
| Genre | Theses |
|---|---|
| Genre | Text |
Bibliographic information
| Statement of responsibility | William Childs. |
|---|---|
| Note | Submitted to the Department of Chemistry. |
| Thesis | Ph. D. Stanford University 2010 |
| Location |
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| OCLC | 747311868 |
| Location | electronic resource |
Access conditions
- Copyright
- © 2010 by William James Childs
- License
- This work is licensed under a Creative Commons Attribution Non Commercial 3.0 Unported license (CC BY-NC).
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