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NMR characterization of intrinsically disordered alpha-synuclein
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Text
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Title
NMR characterization of intrinsically disordered alpha-synuclein
SubTitle
implication for aggregation in Parkinson's disease
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ETD_2300
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http://hdl.rutgers.edu/1782.2/rucore10001600001.ETD.000052165
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eng
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theses
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Chemistry and Chemical Biology
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(authority = ETD-LCSH)
Topic
Parkinsons disease
Subject
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(authority = ETD-LCSH)
Topic
Parkinsons disease--Molecular aspects
Subject
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(authority = ETD-LCSH)
Topic
Protein folding
Subject
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(authority = ETD-LCSH)
Topic
Cell aggregation
Abstract
A broad range of human diseases arise from the loss of the native function of a specific peptide or protein. The pathological conditions of these diseases are generally referred to as protein misfolding and aggregation problem. An increasing number of misfolded proteins are associated with the deposition of protein aggregation in the brain cells resulting in neurodegenerative diseases, for example, the aggregated α-synuclein (αSyn) in sporadic Parkinson’s disease.
αSyn is an intrinsically disordered protein and the aggregation kinetics are sensitive to the changes of amino acids and chemical environments. The structural conversion of αSyn from an unfolded monomeric ensemble to a well-ordered fibril remains unclear. In this work, nuclear magnetic resonance (NMR) was applied to characterize the aggregation-prone species including homologous mouse αSyn and human αSyn at low pH. Comparison of human and mouse αSyn at pH 7.4 and at supercooled aqueous solution presents a less compact mouse αSyn ensemble with transient inter-chain interactions at the N-terminal region. Characterization of αSyn conformational ensembles at both neutral and low pH shows that the altered charge distribution at low pH changes the structural properties of these ensembles and leads to rapid aggregation. Paramagnetic relaxation enhancement (PRE) experiments using mixed isotope (15N) and spin labeled αSyn further illustrate different intermolecular contacts of low-populated transient complex at neutral and low pH. A head-to-tail αSyn complex transiently stabilized by the electrostatic interactions at neutral pH is observed while a tail-to-tail αSyn complex via hydrophobic interactions at low pH is seen. This work provides novel structural insight into the molecular conversion of αSyn at the very early nucleation state.
In addition to understanding the aggregation mechanism of αSyn, an approach that deconvolutes the strong modulation of fast hydrogen exchange (HX) rates on the transverse relaxation rate (R2) of disordered proteins is also presented. This work shows an HX-free R2 profile of αSyn at pH 7.4 and provides an opportunity to understand NMR relaxation of disordered proteins or unstructured loop regions in folded proteins at fast HX conditions such as high pH or high temperature.
αSyn is an intrinsically disordered protein and the aggregation kinetics are sensitive to the changes of amino acids and chemical environments. The structural conversion of αSyn from an unfolded monomeric ensemble to a well-ordered fibril remains unclear. In this work, nuclear magnetic resonance (NMR) was applied to characterize the aggregation-prone species including homologous mouse αSyn and human αSyn at low pH. Comparison of human and mouse αSyn at pH 7.4 and at supercooled aqueous solution presents a less compact mouse αSyn ensemble with transient inter-chain interactions at the N-terminal region. Characterization of αSyn conformational ensembles at both neutral and low pH shows that the altered charge distribution at low pH changes the structural properties of these ensembles and leads to rapid aggregation. Paramagnetic relaxation enhancement (PRE) experiments using mixed isotope (15N) and spin labeled αSyn further illustrate different intermolecular contacts of low-populated transient complex at neutral and low pH. A head-to-tail αSyn complex transiently stabilized by the electrostatic interactions at neutral pH is observed while a tail-to-tail αSyn complex via hydrophobic interactions at low pH is seen. This work provides novel structural insight into the molecular conversion of αSyn at the very early nucleation state.
In addition to understanding the aggregation mechanism of αSyn, an approach that deconvolutes the strong modulation of fast hydrogen exchange (HX) rates on the transverse relaxation rate (R2) of disordered proteins is also presented. This work shows an HX-free R2 profile of αSyn at pH 7.4 and provides an opportunity to understand NMR relaxation of disordered proteins or unstructured loop regions in folded proteins at fast HX conditions such as high pH or high temperature.
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electronic resource
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xv, 200 p. : ill.
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Note
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Ph.D.
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Includes bibliographical references (p. 153-165)
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by Kuen-Phon Wu
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Wu
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Kuen-Phon
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1978-
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Kuen-Phon Wu
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Baum
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Jean
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chair
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Jean Baum
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Levy
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Ronald
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Ronald M Levy
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Talaga
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David
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Advisory Committee
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David S Talaga
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Nanda
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Vikas
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Advisory Committee
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Vikas Nanda
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Rutgers University
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Graduate School - New Brunswick
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2010
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2010-01
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xx
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Rutgers University Electronic Theses and Dissertations
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ETD
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Graduate School - New Brunswick Electronic Theses and Dissertations
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rucore19991600001
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doi:10.7282/T3HT2PGB
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ETD doctoral
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Rights
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The author owns the copyright to this work.
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Copyright protected
Notice
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Open
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Permission or license
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Wu
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Kuen-Phon
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2009-12-14 20:44:07
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Kuen-Phon Wu
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Rutgers University. Graduate School - New Brunswick
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I hereby grant to the Rutgers University Libraries and to my school the non-exclusive right to archive, reproduce and distribute my thesis or dissertation, in whole or in part, and/or my abstract, in whole or in part, in and from an electronic format, subject to the release date subsequently stipulated in this submittal form and approved by my school. I represent and stipulate that the thesis or dissertation and its abstract are my original work, that they do not infringe or violate any rights of others, and that I make these grants as the sole owner of the rights to my thesis or dissertation and its abstract. I represent that I have obtained written permissions, when necessary, from the owner(s) of each third party copyrighted matter to be included in my thesis or dissertation and will supply copies of such upon request by my school. I acknowledge that RU ETD and my school will not distribute my thesis or dissertation or its abstract if, in their reasonable judgment, they believe all such rights have not been secured. I acknowledge that I retain ownership rights to the copyright of my work. I also retain the right to use all or part of this thesis or dissertation in future works, such as articles or books.
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365 days
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