{"id":{"repo_id":"uiuc","oai_identifier":"oai:www.ideals.illinois.edu:2142/101311"},"canonical_url":"https://search.dev.ndltd.org/etd/uiuc/oai:www.ideals.illinois.edu:2142/101311","repository":{"repo_id":"uiuc","name":"University of Illinois - Urbana-Champaign","base_url":"https://www.ideals.illinois.edu/oai-pmh"},"display":{"title":"The effect of single-stranded DNA binding protein RPA2 on XPD helicase processivity","abstract":"Understanding how proteins work together to perform vital cellular functions, such as replicating and repairing DNA, not only extends our understanding of fundamental biology, but can also lead to important medical interventions. As a necessary first step to understanding larger systems, we focus on a two-protein system involved in DNA repair. Xeroderma pigmentosum group D (XPD) is a helicase protein that plays an important role in nucleotide excision repair (NER). Its function is to unwind double-stranded DNA, allowing access to the bases that connect the strands and code genetic information. Previous work has shown that XPD activity is enhanced by the single-stranded DNA binding protein replication protein A (RPA2). However, the mechanism by which unwinding enhancement occurs is unknown. In single-molecule optical trapping experiments, we monitor – with single base-pair precision – the unwinding of a DNA hairpin by XPD in the presence of RPA. We observe the effect of RPA2 on XPD unwinding in real time and distinguish between proposed models of protein cooperation by analyzing changes in unwinding behavior with added RPA. Our data disfavor mechanisms by which RPA2 melts the duplex ahead of XPD as well as RPA2 sequestering ssDNA behind the helicase. We present our own 2-state kinetic model of XPD unwinding that we believe explains our data best. We propose that XPD has two inherent states of unwinding, high and low processivity, and that RPA2 aids unwinding by increasing the likelihood of XPD being in its more processive state.","abstract_html":"Understanding how proteins work together to perform vital cellular functions, such as replicating and repairing DNA, not only extends our understanding of fundamental biology, but can also lead to important medical interventions. As a necessary first step to understanding larger systems, we focus on a two-protein system involved in DNA repair. Xeroderma pigmentosum group D (XPD) is a helicase protein that plays an important role in nucleotide excision repair (NER). Its function is to unwind double-stranded DNA, allowing access to the bases that connect the strands and code genetic information. Previous work has shown that XPD activity is enhanced by the single-stranded DNA binding protein replication protein A (RPA2). However, the mechanism by which unwinding enhancement occurs is unknown. In single-molecule optical trapping experiments, we monitor – with single base-pair precision – the unwinding of a DNA hairpin by XPD in the presence of RPA. We observe the effect of RPA2 on XPD unwinding in real time and distinguish between proposed models of protein cooperation by analyzing changes in unwinding behavior with added RPA. Our data disfavor mechanisms by which RPA2 melts the duplex ahead of XPD as well as RPA2 sequestering ssDNA behind the helicase. We present our own 2-state kinetic model of XPD unwinding that we believe explains our data best. We propose that XPD has two inherent states of unwinding, high and low processivity, and that RPA2 aids unwinding by increasing the likelihood of XPD being in its more processive state.","abstract_has_math":false,"creators":["Stekas, Barbara"],"institution":"University of Illinois at Urbana-Champaign","degree_name":"Ph.D.","degree_level":"Dissertation","degree_discipline":"Physics","degree_department":null,"school":null,"contributors":["Chemla, Yann R.","Aksimentiev, Aleksei","Ha, Taekjip","DeMarco, Brian"],"advisors":[],"committee_chairs":[],"committee_members":[],"year":2018,"date_issued":"2018-09-04T20:47:13Z","date_published":"2018-09-04T20:47:13Z","updated_at":"2026-07-22T22:24:38Z","subjects":["helicase","XPD","RPA","processivity","optical trapping","optical tweezers","nucleotide excision repair","NER"],"languages":["en"],"rights":["Copyright 2018 Barbara Stekas"],"rights_urls":[],"identifier_entries":[]},"links":{"outbound_url":"http://hdl.handle.net/2142/101311","outbound_label":"Handle","outbound_source":"dc:identifier"},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:contributor","label":"Contributor","values":["Chemla, Yann R.","Aksimentiev, Aleksei","Ha, Taekjip","DeMarco, Brian"]},{"key":"dc:creator","label":"Author","values":["Stekas, Barbara"]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"dc:date","label":"Dc Date","values":["2018-09-04T20:47:13Z","2020-09-05T09:15:13Z","2018-04-13","2018-05"]},{"key":"dc:type","label":"Dc Type","values":["text"]},{"key":"thesis:degree_discipline","label":"Discipline","values":["Physics"]},{"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":["helicase","XPD","RPA","processivity","optical trapping","optical tweezers","nucleotide excision repair","NER"]}]},{"id":"language_rights","label":"Language and Rights","entries":[{"key":"dc:language","label":"Dc Language","values":["en"]},{"key":"dc:rights","label":"Dc Rights","values":["Copyright 2018 Barbara Stekas"]}]},{"id":"identifiers","label":"Identifiers","entries":[{"key":"dc:identifier","label":"Identifier","values":["http://hdl.handle.net/2142/101311"]}]},{"id":"additional","label":"Additional Metadata","entries":[{"key":"dc:description","label":"Description","values":["Understanding how proteins work together to perform vital cellular functions, such as replicating and repairing DNA, not only extends our understanding of fundamental biology, but can also lead to important medical interventions. As a necessary first step to understanding larger systems, we focus on a two-protein system involved in DNA repair. Xeroderma pigmentosum group D (XPD) is a helicase protein that plays an important role in nucleotide excision repair (NER). Its function is to unwind double-stranded DNA, allowing access to the bases that connect the strands and code genetic information. Previous work has shown that XPD activity is enhanced by the single-stranded DNA binding protein replication protein A (RPA2). However, the mechanism by which unwinding enhancement occurs is unknown. In single-molecule optical trapping experiments, we monitor – with single base-pair precision – the unwinding of a DNA hairpin by XPD in the presence of RPA. We observe the effect of RPA2 on XPD unwinding in real time and distinguish between proposed models of protein cooperation by analyzing changes in unwinding behavior with added RPA. Our data disfavor mechanisms by which RPA2 melts the duplex ahead of XPD as well as RPA2 sequestering ssDNA behind the helicase. We present our own 2-state kinetic model of XPD unwinding that we believe explains our data best. We propose that XPD has two inherent states of unwinding, high and low processivity, and that RPA2 aids unwinding by increasing the likelihood of XPD being in its more processive state.","Submission published under a 24 month embargo labeled 'Closed Access', the embargo will last until 2020-05-01","The student, Barbara Stekas, accepted the attached license on 2018-04-13 at 11:22.","The student, Barbara Stekas, submitted this Dissertation for approval on 2018-04-13 at 14:36.","This Dissertation was approved for publication on 2018-04-13 at 17:08.","DSpace SAF Submission Ingestion Package generated from Vireo submission #12233 on 2018-08-31 at 17:28:49","Made available in DSpace on 2018-09-04T20:47:13Z (GMT). No. of bitstreams: 4 STEKAS-DISSERTATION-2018.pdf: 12996179 bytes, checksum: 939f04530253bf54cde8e1053a811d3f (MD5) LICENSE.txt: 4211 bytes, checksum: 29332fafe3fc5ec02bf8c5554e484053 (MD5) PROQUEST_LICENSE.txt: 4557 bytes, checksum: 67758a533d9bfdb2d9d080439eada4f2 (MD5) RightsLink-Order.pdf: 67391 bytes, checksum: 288bf3b5e0d7946feb349ee1d79e426a (MD5) Previous issue date: 2018-04-13","Embargo set by: Seth Robbins for item 107396 Lift date: 2020-09-04T20:47:38Z Reason: Author requested closed access (OA after 2yrs) in Vireo ETD system","Embargo set by: Seth Robbins for item 107396 Lift date: 2020-09-04T20:50:11Z Reason: Author requested closed access (OA after 2yrs) in Vireo ETD system","Limited Restriction Lifted for Item 107396 on 2020-09-05T09:15:13Z."]},{"key":"dc:format","label":"Dc Format","values":["application/pdf"]},{"key":"dc:title","label":"Title","values":["The effect of single-stranded DNA binding protein RPA2 on XPD helicase processivity"]}]}],"canonical_facts":{"dc:contributor":["Chemla, Yann R.","Aksimentiev, Aleksei","Ha, Taekjip","DeMarco, Brian"],"dc:creator":["Stekas, Barbara"],"dc:date":["2018-09-04T20:47:13Z","2020-09-05T09:15:13Z","2018-04-13","2018-05"],"dc:description":["Understanding how proteins work together to perform vital cellular functions, such as replicating and repairing DNA, not only extends our understanding of fundamental biology, but can also lead to important medical interventions. As a necessary first step to understanding larger systems, we focus on a two-protein system involved in DNA repair. Xeroderma pigmentosum group D (XPD) is a helicase protein that plays an important role in nucleotide excision repair (NER). Its function is to unwind double-stranded DNA, allowing access to the bases that connect the strands and code genetic information. Previous work has shown that XPD activity is enhanced by the single-stranded DNA binding protein replication protein A (RPA2). However, the mechanism by which unwinding enhancement occurs is unknown. In single-molecule optical trapping experiments, we monitor – with single base-pair precision – the unwinding of a DNA hairpin by XPD in the presence of RPA. We observe the effect of RPA2 on XPD unwinding in real time and distinguish between proposed models of protein cooperation by analyzing changes in unwinding behavior with added RPA. Our data disfavor mechanisms by which RPA2 melts the duplex ahead of XPD as well as RPA2 sequestering ssDNA behind the helicase. We present our own 2-state kinetic model of XPD unwinding that we believe explains our data best. We propose that XPD has two inherent states of unwinding, high and low processivity, and that RPA2 aids unwinding by increasing the likelihood of XPD being in its more processive state.","Submission published under a 24 month embargo labeled 'Closed Access', the embargo will last until 2020-05-01","The student, Barbara Stekas, accepted the attached license on 2018-04-13 at 11:22.","The student, Barbara Stekas, submitted this Dissertation for approval on 2018-04-13 at 14:36.","This Dissertation was approved for publication on 2018-04-13 at 17:08.","DSpace SAF Submission Ingestion Package generated from Vireo submission #12233 on 2018-08-31 at 17:28:49","Made available in DSpace on 2018-09-04T20:47:13Z (GMT). No. of bitstreams: 4 STEKAS-DISSERTATION-2018.pdf: 12996179 bytes, checksum: 939f04530253bf54cde8e1053a811d3f (MD5) LICENSE.txt: 4211 bytes, checksum: 29332fafe3fc5ec02bf8c5554e484053 (MD5) PROQUEST_LICENSE.txt: 4557 bytes, checksum: 67758a533d9bfdb2d9d080439eada4f2 (MD5) RightsLink-Order.pdf: 67391 bytes, checksum: 288bf3b5e0d7946feb349ee1d79e426a (MD5) Previous issue date: 2018-04-13","Embargo set by: Seth Robbins for item 107396 Lift date: 2020-09-04T20:47:38Z Reason: Author requested closed access (OA after 2yrs) in Vireo ETD system","Embargo set by: Seth Robbins for item 107396 Lift date: 2020-09-04T20:50:11Z Reason: Author requested closed access (OA after 2yrs) in Vireo ETD system","Limited Restriction Lifted for Item 107396 on 2020-09-05T09:15:13Z."],"dc:format":["application/pdf"],"dc:identifier":["http://hdl.handle.net/2142/101311"],"dc:language":["en"],"dc:rights":["Copyright 2018 Barbara Stekas"],"dc:subject":["helicase","XPD","RPA","processivity","optical trapping","optical tweezers","nucleotide excision repair","NER"],"dc:title":["The effect of single-stranded DNA binding protein RPA2 on XPD helicase processivity"],"dc:type":["text"],"thesis:degree_discipline":["Physics"],"thesis:degree_level":["Dissertation"],"thesis:degree_name":["Ph.D."],"thesis:institution_name":["University of Illinois at Urbana-Champaign"]},"updated_at":"2026-07-22T22:24:38Z"}