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Biophysical Studies of Ligand-gated Ion Channels

Citation

Wong, Betty Ko (2017) Biophysical Studies of Ligand-gated Ion Channels. Dissertation (Ph.D.), California Institute of Technology. doi:10.7907/Z9TT4P0Q. http://resolver.caltech.edu/CaltechTHESIS:05302017-225018783

Abstract

This dissertation describes building a methodology for and the biophysical studies of ligand-gated ion channels (LGICs).

The primary focus of the first half of this dissertation is on developing a fluorescence-based assay to broadly study LGICs. Chapter 2 describes the site-selective incorporation of a turn-on fluorophore via unnatural amino acid mutagenesis on the mouse muscle-type nicotinic acetylcholine receptor (nAChR) in Xenopus oocytes as a proof-of-principle study. This method has proven to yield very low levels of undesired fluorescent background, which was a problem for previous incorporation techniques. Chapter 3 describes efforts towards imaging this in vivo system using lifetime imaging with efforts hampered by the inability to detect a clear signal. Chapter 4 describes efforts to apply the lifetime imaging approach towards a different system involving 5-HT3 proteins fused to fluorescent proteins in COS-7 cells.

The second half of this dissertation focuses on studies of menthol, a flavorant added to cigarettes that contributes to smoking addiction, as a negative allosteric modulator of the αβ42 nAChR. Chapter 5 reveals the stereochemical effects, or rather lack of, of menthol on the two stoichiometries of the αβ42 receptor. Chapter 6 seeks to identify the residue interactions with menthol of the αβ42 receptor using a combination of computational and experimental studies.

Item Type:Thesis (Dissertation (Ph.D.))
Subject Keywords:ion channel; neuroscience; nicotinic acetylcholine receptor; non-canonical amino acid; menthol; fluorescence microscopy
Degree Grantor:California Institute of Technology
Division:Chemistry and Chemical Engineering
Major Option:Biochemistry and Molecular Biophysics
Thesis Availability:Public (worldwide access)
Research Advisor(s):
  • Dougherty, Dennis A. (advisor)
  • Lester, Henry A. (co-advisor)
Thesis Committee:
  • Gradinaru, Viviana
  • Rees, Douglas C. (chair)
  • Dougherty, Dennis A.
  • Lester, Henry A.
Defense Date:22 May 2017
Non-Caltech Author Email:bwcaltech (AT) gmail.com
Funders:
Funding AgencyGrant Number
National Science Foundation Graduate Research FellowshipUNSPECIFIED
Record Number:CaltechTHESIS:05302017-225018783
Persistent URL:http://resolver.caltech.edu/CaltechTHESIS:05302017-225018783
DOI:10.7907/Z9TT4P0Q
Default Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:10227
Collection:CaltechTHESIS
Deposited By: Betty Wong
Deposited On:06 Jun 2017 23:40
Last Modified:08 Jun 2018 20:14

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