{"id":{"repo_id":"uiuc","oai_identifier":"oai:www.ideals.illinois.edu:2142/122222"},"canonical_url":"https://search.dev.ndltd.org/etd/uiuc/oai:www.ideals.illinois.edu:2142/122222","repository":{"repo_id":"uiuc","name":"University of Illinois - Urbana-Champaign","base_url":"https://www.ideals.illinois.edu/oai-pmh"},"display":{"title":"Development and application of quantitative ligand-binding studies to elucidate mechanistic aspects of pentameric ligand-gated ion channels","abstract":"Submission published under a 24 month embargo labeled 'Closed Access', the embargo will last until 2025-12-01","abstract_html":"Submission published under a 24 month embargo labeled 'Closed Access', the embargo will last until 2025-12-01","abstract_has_math":false,"creators":["Godellas, Nicole E"],"institution":"University of Illinois at Urbana-Champaign","degree_name":"Ph.D.","degree_level":"Dissertation","degree_discipline":"Molecular & Integrative Physi","degree_department":null,"school":null,"contributors":["Grosman, Claudio","Anakk, Sayeepriyadarshini","Llano, Daniel A","Lingle, Christopher J"],"advisors":[],"committee_chairs":[],"committee_members":[],"year":2023,"date_issued":"2023-12","date_published":"2023-12","updated_at":"2026-07-22T22:25:00Z","subjects":["Ligand-binding Assays","Cys-loop Receptors","Pentameric Ligand-gated Ion Channels","Function","Nicotinic Receptors","Ion Channels","Mechanisms"],"languages":["en","eng"],"rights":["Copyright 2023 Nicole Godellas"],"rights_urls":[],"identifier_entries":[]},"links":{"outbound_url":"https://hdl.handle.net/2142/122222","outbound_label":"Handle","outbound_source":"dc:identifier"},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:contributor","label":"Contributor","values":["Grosman, Claudio","Anakk, Sayeepriyadarshini","Llano, Daniel A","Lingle, Christopher J"]},{"key":"dc:creator","label":"Author","values":["Godellas, Nicole E"]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"dc:date","label":"Dc Date","values":["2023-12","2023-11-29"]},{"key":"dc:type","label":"Dc Type","values":["text"]},{"key":"thesis:degree_discipline","label":"Discipline","values":["Molecular & Integrative Physi"]},{"key":"thesis:degree_level","label":"Degree Level","values":["Dissertation"]},{"key":"thesis:degree_name","label":"Degree Name","values":["Ph.D."]},{"key":"thesis:institution_name","label":"Thesis Institution Name","values":["University of Illinois at Urbana-Champaign"]}]},{"id":"subjects_keywords","label":"Subjects and Keywords","entries":[{"key":"dc:subject","label":"Dc Subject","values":["Ligand-binding Assays","Cys-loop Receptors","Pentameric Ligand-gated Ion Channels","Function","Nicotinic Receptors","Ion Channels","Mechanisms"]}]},{"id":"language_rights","label":"Language and Rights","entries":[{"key":"dc:language","label":"Dc Language","values":["en","eng"]},{"key":"dc:rights","label":"Dc Rights","values":["Copyright 2023 Nicole Godellas"]}]},{"id":"identifiers","label":"Identifiers","entries":[{"key":"dc:identifier","label":"Identifier","values":["https://hdl.handle.net/2142/122222"]}]},{"id":"additional","label":"Additional Metadata","entries":[{"key":"dc:description","label":"Description","values":["Submission published under a 24 month embargo labeled 'Closed Access', the embargo will last until 2025-12-01","The student, Nicole Godellas, accepted the attached license on 2023-11-15 at 16:38.","The student, Nicole Godellas, submitted this Dissertation for approval on 2023-11-15 at 17:07.","This Dissertation was approved for publication on 2023-11-29 at 11:52.","DSpace SAF Submission Ingestion Package generated from Vireo submission #19932 on 2024-03-01 at 13:48:28","Mechanistic studies of ligand-gated ion channels are not only an indispensable component of our understanding of Biology, but also, a fundamental step for the rational design of targeted therapeutics. For several decades, ion-channel mechanisms have been elucidated by means of electrophysiological studies of mutants; however, studying the effect of loss-of-function mutations (that is, mutations that render the channel “electrically silent”) requires an alternative method. Here, we have developed, optimized, and applied an equilibrium-type ligand-binding assay to a number of questions in the field of ion-channel physiology that cannot be answered by measuring ion transport. The application of ligand-binding assays to probe function in ion channels is, by no means, new; this classical assay emerged decades ago. However, its application to the mechanistic study of receptor-channels received little attention. We began with both a practical and theoretical study of this approach and optimized the methodological conditions for a competition ligand-binding assay using the human homomeric α7 AChR, radiolabeled α-bungarotoxin (α-BgTx), unlabeled small-molecule cholinergic ligands, and calculations in the framework of a kinetic reaction scheme modeling our pentameric receptor of interest. We, then, applied this approach to the elucidation of mechanistic aspects of these ligand-gated ion channels. We concluded that: 1) Ligand-binding affinities are insensitive to binding-site occupancy; 2) Mutations that are distant from the orthosteric-binding sites (say, in the transmembrane domain) have little to no effect on the channel’s affinity for orthosteric ligand; 3) The binding of the SARS-CoV-2 spike protein to the orthosteric-binding sites of the human α7 nAChR (and its subsequent competition with acetylcholine, choline, or nicotine) is unlikely to be a relevant aspect of this disease; and 4) The distance between the extracellular (ligand-binding) domain and the transmembrane-pore domain is critical for effective binding–gating coupling."]},{"key":"dc:format","label":"Dc Format","values":["application/pdf"]},{"key":"dc:title","label":"Title","values":["Development and application of quantitative ligand-binding studies to elucidate mechanistic aspects of pentameric ligand-gated ion channels"]}]}],"canonical_facts":{"dc:contributor":["Grosman, Claudio","Anakk, Sayeepriyadarshini","Llano, Daniel A","Lingle, Christopher J"],"dc:creator":["Godellas, Nicole E"],"dc:date":["2023-12","2023-11-29"],"dc:description":["Submission published under a 24 month embargo labeled 'Closed Access', the embargo will last until 2025-12-01","The student, Nicole Godellas, accepted the attached license on 2023-11-15 at 16:38.","The student, Nicole Godellas, submitted this Dissertation for approval on 2023-11-15 at 17:07.","This Dissertation was approved for publication on 2023-11-29 at 11:52.","DSpace SAF Submission Ingestion Package generated from Vireo submission #19932 on 2024-03-01 at 13:48:28","Mechanistic studies of ligand-gated ion channels are not only an indispensable component of our understanding of Biology, but also, a fundamental step for the rational design of targeted therapeutics. For several decades, ion-channel mechanisms have been elucidated by means of electrophysiological studies of mutants; however, studying the effect of loss-of-function mutations (that is, mutations that render the channel “electrically silent”) requires an alternative method. Here, we have developed, optimized, and applied an equilibrium-type ligand-binding assay to a number of questions in the field of ion-channel physiology that cannot be answered by measuring ion transport. The application of ligand-binding assays to probe function in ion channels is, by no means, new; this classical assay emerged decades ago. However, its application to the mechanistic study of receptor-channels received little attention. We began with both a practical and theoretical study of this approach and optimized the methodological conditions for a competition ligand-binding assay using the human homomeric α7 AChR, radiolabeled α-bungarotoxin (α-BgTx), unlabeled small-molecule cholinergic ligands, and calculations in the framework of a kinetic reaction scheme modeling our pentameric receptor of interest. We, then, applied this approach to the elucidation of mechanistic aspects of these ligand-gated ion channels. We concluded that: 1) Ligand-binding affinities are insensitive to binding-site occupancy; 2) Mutations that are distant from the orthosteric-binding sites (say, in the transmembrane domain) have little to no effect on the channel’s affinity for orthosteric ligand; 3) The binding of the SARS-CoV-2 spike protein to the orthosteric-binding sites of the human α7 nAChR (and its subsequent competition with acetylcholine, choline, or nicotine) is unlikely to be a relevant aspect of this disease; and 4) The distance between the extracellular (ligand-binding) domain and the transmembrane-pore domain is critical for effective binding–gating coupling."],"dc:format":["application/pdf"],"dc:identifier":["https://hdl.handle.net/2142/122222"],"dc:language":["en","eng"],"dc:rights":["Copyright 2023 Nicole Godellas"],"dc:subject":["Ligand-binding Assays","Cys-loop Receptors","Pentameric Ligand-gated Ion Channels","Function","Nicotinic Receptors","Ion Channels","Mechanisms"],"dc:title":["Development and application of quantitative ligand-binding studies to elucidate mechanistic aspects of pentameric ligand-gated ion channels"],"dc:type":["text"],"thesis:degree_discipline":["Molecular & Integrative Physi"],"thesis:degree_level":["Dissertation"],"thesis:degree_name":["Ph.D."],"thesis:institution_name":["University of Illinois at Urbana-Champaign"]},"updated_at":"2026-07-22T22:25:00Z"}