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Structure-Function Studies of Nicotinic Acetylcholine Receptors Using Selective Agonists and Positive Allosteric Modulators

Citation

Marotta, Christopher Bruno (2015) Structure-Function Studies of Nicotinic Acetylcholine Receptors Using Selective Agonists and Positive Allosteric Modulators. Dissertation (Ph.D.), California Institute of Technology. doi:10.7907/Z9V122Q9. https://resolver.caltech.edu/CaltechTHESIS:05292015-144036736

Abstract

This dissertation primarily describes chemical-scale studies of nicotinic acetylcholine receptors (nAChRs) in order to better understand ligand-receptor selectivity and allosteric modulation influences during receptor activation. Electrophysiology coupled with canonical and non-canonical amino acids mutagenesis is used to probe subtle changes in receptor function.

The first half of this dissertation focuses on differential agonist selectivity of α4β2-containing nAChRs. The α4β2 nAChR can assemble in alternative stoichiometries as well as assemble with other accessory subunits. Chapter 2 identifies key structural residues that dictate binding and activation of three stoichiometry-dependent α4β2 receptor ligands: sazetidine-A, cytisine, and NS9283. These do not follow previously suggested hydrogen-bonding patterns of selectivity. Instead, three residues on the complementary subunit strongly influence binding ability of a ligand and receptor activation. Chapter 3 involves isolation of a α5α4β2 receptor-enriched population to test for a potential alternative agonist binding location at the α5 α4 interface. Results strongly suggest that agonist occupation of this site is not necessary for receptor activation and that the α5 subunit only incorporates at the accessory subunit location.

The second half of this dissertation seeks to identify residue interactions with positive allosteric modulators (PAMs) of the α7 nAChR. Chapter 4 focuses on methods development to study loss of potentiation of Type I PAMs, which indicate residues vital to propagation of PAM effects and/or binding. Chapter 5 investigates α7 receptor modulation by a Type II PAM (PNU 120596). These results show that PNU 120596 does not alter the agonist binding site, thus is relegated to influencing only the gating component of activation. From this, we were able to map a potential network of residues from the agonist binding site to the proposed PNU 120596 binding site that are essential for receptor potentiation.

Item Type:Thesis (Dissertation (Ph.D.))
Subject Keywords:nicotinic acetylcholine receptors; non-canonical amino acid; structure-function; positive allosteric modulator; selective agonists
Degree Grantor:California Institute of Technology
Division:Chemistry and Chemical Engineering
Major Option:Chemistry
Thesis Availability:Public (worldwide access)
Research Advisor(s):
  • Dougherty, Dennis A.
Thesis Committee:
  • Rees, Douglas C. (chair)
  • Barton, Jacqueline K.
  • Miller, Thomas F.
  • Dougherty, Dennis A.
Defense Date:22 May 2015
Funders:
Funding AgencyGrant Number
NIHNS 34407
NIHDA017279
NIHDA280382
NIH/NRSAGM07616
California Tobacco-Related Disease Research Program from the UNiveristy of California19XT-0102
Record Number:CaltechTHESIS:05292015-144036736
Persistent URL:https://resolver.caltech.edu/CaltechTHESIS:05292015-144036736
DOI:10.7907/Z9V122Q9
Default Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:8927
Collection:CaltechTHESIS
Deposited By: Christopher Marotta
Deposited On:02 Jun 2015 15:26
Last Modified:04 Oct 2019 00:08

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