Developing a general description of the unfolded state of a protein
This thesis focuses on both the dynamical and structural aspects of the unfolded states of a protein and develops a general description of these unfolded states over all denaturing conditions. Investigating the unfolded states under physiological conditions can provide insights into the entire folding pathway as they define the most likely conformations before the folding transition. Also, since the initial motion of a protein in solvent early in the folding process is driven by diffusion, the intramolecular diffusion rate may set an upper speed limit for protein folding.In this thesis, the experimental measurement of the intramolecular diffusion is achieved by a pump-probe spectroscopy using the technique of Trp/Cys contact quenching. Three typical proteins in different folding categories have been studied: the intrinsically disordered apocytochrome C, the aggregation-prone HypF-N and the well-folded ACBP (Acyl-coenzyme A-binding Protein). For ACBP, I also used a microfluidic mixer coupled with the pump-probe spectroscopy to study the intramolecular loop contact formation in its refolding process. In order to extract the effective diffusion coefficient from the observed contact rates, I employed a polymer theory by Szabo, Schulten and Schulten (SSS) which requires a probability distribution of equilibrium Trp/Cys distances. In an attempt to capture the transient residual structures and intramolecular interactions of unfolded polypeptides under folding conditions, I integrated sequence specificities into a traditional non-overlapping worm-like chain model. The new model can statistically re-weight the distribution to favor those conformations with more hydrophobic contacts and it yields quite convergent results over many tested sequences. In the thesis, I will also show that this model quantitatively predicted paramagnetic resonance enhancement (PRE) measurements of ACBP and DrkN with no adjustable parameters.After applying the SSS theory with the Trp/Cys distance distributions produced by the energy re-weighted worm-like chain model to calculating the intramolecular diffusion coefficients of those experimentally studied proteins, I found an intrinsic relationship between the intramolecular diffusion rates and the aggregation propensities and I propose a dynamic range of conformational reorganization within which partially or fully unfolded states are prone to aggregation.
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- In Collections
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Electronic Theses & Dissertations
- Copyright Status
- In Copyright
- Material Type
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Theses
- Authors
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Chen, Yujie
- Thesis Advisors
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Lapidus, Lisa
- Committee Members
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Lai, Chih-Wei
Feig, Michael
Duxbury, Phillip
Tessmer, Stuart
- Date
- 2011
- Subjects
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Protein folding
Proteins
- Program of Study
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Physics
- Degree Level
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Doctoral
- Language
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English
- Pages
- xii, 107 pages
- ISBN
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9781124554044
1124554041
- Permalink
- https://doi.org/doi:10.25335/8rx3-6q52