{"id":{"repo_id":"uiuc","oai_identifier":"oai:www.ideals.illinois.edu:2142/88273"},"canonical_url":"https://search.dev.ndltd.org/etd/uiuc/oai:www.ideals.illinois.edu:2142/88273","repository":{"repo_id":"uiuc","name":"University of Illinois - Urbana-Champaign","base_url":"https://www.ideals.illinois.edu/oai-pmh"},"display":{"title":"Probing surface protein patterning in biological systems using fluorescence nanoscopy","abstract":"A complete understanding of biological substructures is often obscured by the diffraction limit of visible light in conventional fluorescence microscopy. Recently developed fluorescence nanoscopy techniques such as stochastic optical reconstruction microscopy (STORM) effectively break the diffraction limit (~250 nm) to enable imaging with an order of magnitude smaller than the spatial resolution. In this work, we applied fluorescence nanoscopy to study the patterning of proteins in biological surfaces at a high spatial resolution of 25 nm. We also developed new analytical methods to extract quantitative information on protein arrangement within biological substrates to elucidate key phenomenon such as signal amplification in bacteria during chemotaxis. In B. subtilis, we observed reorganization in arrays of McpB receptor proteins, key chemotactic receptors for asparagine sensing, upon exposure to saturating stimulant concentrations. More specifically, receptors formed large polar clusters in the absence of stimulant but shifted towards smaller and more dispersed lattices throughout the cell when stimulant was added. In a first-of-its-kind fluorescence nanoscopic study on B. subtilis, we measured cluster sizes and intra cluster density of these receptors. Our results support the existing theories on the role of cooperativity via receptor clustering in signal amplification to respond to wide range of external stimuli in bacteria. In second project, we studied the role of viral proteins on HIV-1 infectivity. In HIV-1, envelope proteins gp120 and gp41 are known to play a significant role in infectivity and their arrangement is indicative of internal viral structure at different stages of its life cycle. We ascertained the distribution of both the envelope proteins on surface of virions at a high resolution of ~25 nm using three-color fluorescence nanoscopy. Our work effectively paves the way for revealing new levels of organization of surface proteins at nanometer scale spatial resolution for a molecular view understanding of dynamics in biological systems.","abstract_html":"A complete understanding of biological substructures is often obscured by the diffraction limit of visible light in conventional fluorescence microscopy. Recently developed fluorescence nanoscopy techniques such as stochastic optical reconstruction microscopy (STORM) effectively break the diffraction limit (~250 nm) to enable imaging with an order of magnitude smaller than the spatial resolution. In this work, we applied fluorescence nanoscopy to study the patterning of proteins in biological surfaces at a high spatial resolution of 25 nm. We also developed new analytical methods to extract quantitative information on protein arrangement within biological substrates to elucidate key phenomenon such as signal amplification in bacteria during chemotaxis. In B. subtilis, we observed reorganization in arrays of McpB receptor proteins, key chemotactic receptors for asparagine sensing, upon exposure to saturating stimulant concentrations. More specifically, receptors formed large polar clusters in the absence of stimulant but shifted towards smaller and more dispersed lattices throughout the cell when stimulant was added. In a first-of-its-kind fluorescence nanoscopic study on B. subtilis, we measured cluster sizes and intra cluster density of these receptors. Our results support the existing theories on the role of cooperativity via receptor clustering in signal amplification to respond to wide range of external stimuli in bacteria. In second project, we studied the role of viral proteins on HIV-1 infectivity. In HIV-1, envelope proteins gp120 and gp41 are known to play a significant role in infectivity and their arrangement is indicative of internal viral structure at different stages of its life cycle. We ascertained the distribution of both the envelope proteins on surface of virions at a high resolution of ~25 nm using three-color fluorescence nanoscopy. Our work effectively paves the way for revealing new levels of organization of surface proteins at nanometer scale spatial resolution for a molecular view understanding of dynamics in biological systems.","abstract_has_math":false,"creators":["Agrawal, Utsav"],"institution":"University of Illinois at Urbana-Champaign","degree_name":"Ph.D.","degree_level":"Dissertation","degree_discipline":"Chemical Engineering","degree_department":null,"school":null,"contributors":["Schroeder, Charles M.","Kraft, Mary L.","Rao, Christopher V.","Kuhlman, Thomas E."],"advisors":[],"committee_chairs":[],"committee_members":[],"year":2015,"date_issued":"2015-09-29T21:03:10Z","date_published":"2015-09-29T21:03:10Z","updated_at":"2026-07-22T22:26:31Z","subjects":["fluorescence nanoscopy","stochastic optical reconstruction microscopy (STORM)","B. subtilis","human immunodeficiency virus (HIV)","chemotaxis","McpB","envelope proteins","clustering"],"languages":["en"],"rights":["Copyright 2015 Utsav Agrawal"],"rights_urls":[],"identifier_entries":[]},"links":{"outbound_url":"http://hdl.handle.net/2142/88273","outbound_label":"Handle","outbound_source":"dc:identifier"},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:contributor","label":"Contributor","values":["Schroeder, Charles M.","Kraft, Mary L.","Rao, Christopher V.","Kuhlman, Thomas E."]},{"key":"dc:creator","label":"Author","values":["Agrawal, Utsav"]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"dc:date","label":"Dc Date","values":["2015-09-29T21:03:10Z","2017-09-30T09:15:30Z","2015-08","2015-07-10","2015-8"]},{"key":"dc:type","label":"Dc Type","values":["text"]},{"key":"thesis:degree_discipline","label":"Discipline","values":["Chemical Engineering"]},{"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":["fluorescence nanoscopy","stochastic optical reconstruction microscopy (STORM)","B. subtilis","human immunodeficiency virus (HIV)","chemotaxis","McpB","envelope proteins","clustering"]}]},{"id":"language_rights","label":"Language and Rights","entries":[{"key":"dc:language","label":"Dc Language","values":["en"]},{"key":"dc:rights","label":"Dc Rights","values":["Copyright 2015 Utsav Agrawal"]}]},{"id":"identifiers","label":"Identifiers","entries":[{"key":"dc:identifier","label":"Identifier","values":["http://hdl.handle.net/2142/88273"]}]},{"id":"additional","label":"Additional Metadata","entries":[{"key":"dc:description","label":"Description","values":["A complete understanding of biological substructures is often obscured by the diffraction limit of visible light in conventional fluorescence microscopy. Recently developed fluorescence nanoscopy techniques such as stochastic optical reconstruction microscopy (STORM) effectively break the diffraction limit (~250 nm) to enable imaging with an order of magnitude smaller than the spatial resolution. In this work, we applied fluorescence nanoscopy to study the patterning of proteins in biological surfaces at a high spatial resolution of 25 nm. We also developed new analytical methods to extract quantitative information on protein arrangement within biological substrates to elucidate key phenomenon such as signal amplification in bacteria during chemotaxis. In B. subtilis, we observed reorganization in arrays of McpB receptor proteins, key chemotactic receptors for asparagine sensing, upon exposure to saturating stimulant concentrations. More specifically, receptors formed large polar clusters in the absence of stimulant but shifted towards smaller and more dispersed lattices throughout the cell when stimulant was added. In a first-of-its-kind fluorescence nanoscopic study on B. subtilis, we measured cluster sizes and intra cluster density of these receptors. Our results support the existing theories on the role of cooperativity via receptor clustering in signal amplification to respond to wide range of external stimuli in bacteria. In second project, we studied the role of viral proteins on HIV-1 infectivity. In HIV-1, envelope proteins gp120 and gp41 are known to play a significant role in infectivity and their arrangement is indicative of internal viral structure at different stages of its life cycle. We ascertained the distribution of both the envelope proteins on surface of virions at a high resolution of ~25 nm using three-color fluorescence nanoscopy. Our work effectively paves the way for revealing new levels of organization of surface proteins at nanometer scale spatial resolution for a molecular view understanding of dynamics in biological systems.","Submission published under a 24 month embargo labeled 'Closed Access', the embargo will last until 2017-08-01","The student, Utsav Agrawal, accepted the attached license on 2015-07-09 at 15:02.","The student, Utsav Agrawal, submitted this Dissertation for approval on 2015-07-09 at 15:03.","This Dissertation was approved for publication on 2015-07-10 at 08:51.","DSpace SAF Submission Ingestion Package generated from Vireo submission #8391 on 2015-09-29 at 15:05:53","Made available in DSpace on 2015-09-29T21:03:10Z (GMT). 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Recently developed fluorescence nanoscopy techniques such as stochastic optical reconstruction microscopy (STORM) effectively break the diffraction limit (~250 nm) to enable imaging with an order of magnitude smaller than the spatial resolution. In this work, we applied fluorescence nanoscopy to study the patterning of proteins in biological surfaces at a high spatial resolution of 25 nm. We also developed new analytical methods to extract quantitative information on protein arrangement within biological substrates to elucidate key phenomenon such as signal amplification in bacteria during chemotaxis. In B. subtilis, we observed reorganization in arrays of McpB receptor proteins, key chemotactic receptors for asparagine sensing, upon exposure to saturating stimulant concentrations. More specifically, receptors formed large polar clusters in the absence of stimulant but shifted towards smaller and more dispersed lattices throughout the cell when stimulant was added. In a first-of-its-kind fluorescence nanoscopic study on B. subtilis, we measured cluster sizes and intra cluster density of these receptors. Our results support the existing theories on the role of cooperativity via receptor clustering in signal amplification to respond to wide range of external stimuli in bacteria. In second project, we studied the role of viral proteins on HIV-1 infectivity. In HIV-1, envelope proteins gp120 and gp41 are known to play a significant role in infectivity and their arrangement is indicative of internal viral structure at different stages of its life cycle. We ascertained the distribution of both the envelope proteins on surface of virions at a high resolution of ~25 nm using three-color fluorescence nanoscopy. Our work effectively paves the way for revealing new levels of organization of surface proteins at nanometer scale spatial resolution for a molecular view understanding of dynamics in biological systems.","Submission published under a 24 month embargo labeled 'Closed Access', the embargo will last until 2017-08-01","The student, Utsav Agrawal, accepted the attached license on 2015-07-09 at 15:02.","The student, Utsav Agrawal, submitted this Dissertation for approval on 2015-07-09 at 15:03.","This Dissertation was approved for publication on 2015-07-10 at 08:51.","DSpace SAF Submission Ingestion Package generated from Vireo submission #8391 on 2015-09-29 at 15:05:53","Made available in DSpace on 2015-09-29T21:03:10Z (GMT). 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