{"id":{"repo_id":"uiuc","oai_identifier":"oai:www.ideals.illinois.edu:2142/20509"},"canonical_url":"https://search.dev.ndltd.org/etd/uiuc/oai:www.ideals.illinois.edu:2142/20509","repository":{"repo_id":"uiuc","name":"University of Illinois - Urbana-Champaign","base_url":"https://www.ideals.illinois.edu/oai-pmh"},"display":{"title":"Macromolecular recognition in the cytochrome P450(cam) enzyme system","abstract":"The cytochrome P-450$\\sb{\\rm cam}$ reaction cycle is a complex set of coordinated chemical transformations requiring precise temporal and spatial control of reactivities. Cytochrome P-450$\\sb{\\rm cam}$ catalyzes the regio- and stereo-specific hydroxylation of camphor to form 5-exo-hydroxycamphor. The two reducing equivalents required for this reaction are supplied physiologically by putidaredoxin, a Fe$\\sb2$S$\\sb2$ iron-sulfur protein. The mammalian cytochromes P-450 are also known to interact with cytochrome b$\\sb5$, a small redox protein for which a high resolution crystal structure is available. To characterize the molecular cytochrome P-450$\\sb{\\rm cam}$ binding surface, cytochrome b$\\sb5$ was first genetically engineered to afford a fluorescent derivative capable of monitoring its association with cytochrome P-450$\\sb{\\rm cam}$. The interaction was subsequently computer modeled by looking for van der Waals complementarity and salt bridge formation between the cytochrome b$\\sb5$ anionic binding surface and basic residues on the cytochrome P-450$\\sb{\\rm cam}$ surface. A good fit was found on the proximal surface of nearest approach to the cytochrome P-450$\\sb{\\rm cam}$ heme prosthetic group.","abstract_html":"The cytochrome P-450$\\sb{\\rm cam}$ reaction cycle is a complex set of coordinated chemical transformations requiring precise temporal and spatial control of reactivities. Cytochrome P-450$\\sb{\\rm cam}$ catalyzes the regio- and stereo-specific hydroxylation of camphor to form 5-exo-hydroxycamphor. The two reducing equivalents required for this reaction are supplied physiologically by putidaredoxin, a Fe$\\sb2$S$\\sb2$ iron-sulfur protein. The mammalian cytochromes P-450 are also known to interact with cytochrome b$\\sb5$, a small redox protein for which a high resolution crystal structure is available. To characterize the molecular cytochrome P-450$\\sb{\\rm cam}$ binding surface, cytochrome b$\\sb5$ was first genetically engineered to afford a fluorescent derivative capable of monitoring its association with cytochrome P-450$\\sb{\\rm cam}$. The interaction was subsequently computer modeled by looking for van der Waals complementarity and salt bridge formation between the cytochrome b$\\sb5$ anionic binding surface and basic residues on the cytochrome P-450$\\sb{\\rm cam}$ surface. A good fit was found on the proximal surface of nearest approach to the cytochrome P-450$\\sb{\\rm cam}$ heme prosthetic group.","abstract_has_math":true,"creators":["Stayton, Patrick Sean"],"institution":"University of Illinois at Urbana-Champaign","degree_name":"Ph.D.","degree_level":"Dissertation","degree_discipline":"Biochemistry","degree_department":null,"school":null,"contributors":["Sligar, Stephen G."],"advisors":[],"committee_chairs":[],"committee_members":[],"year":2011,"date_issued":"2011-05-07T12:41:15Z","date_published":"2011-05-07T12:41:15Z","updated_at":"2026-07-22T22:25:16Z","subjects":["Chemistry, Biochemistry","Biophysics, General"],"languages":["eng"],"rights":["Copyright 1989 Stayton, Patrick Sean"],"rights_urls":[],"identifier_entries":[{"key":"dc:identifier","label":"Identifier","values":["AAI9011038","(UMI)AAI9011038"],"render_values":[{"text":"AAI9011038","href":null,"code":true},{"text":"(UMI)AAI9011038","href":null,"code":true}]}]},"links":{"outbound_url":"http://hdl.handle.net/2142/20509","outbound_label":"Handle","outbound_source":"dc:identifier"},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:contributor","label":"Contributor","values":["Sligar, Stephen G."]},{"key":"dc:creator","label":"Author","values":["Stayton, Patrick Sean"]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"dc:date","label":"Dc Date","values":["2011-05-07T12:41:15Z","10000-01-01","1989"]},{"key":"dc:type","label":"Dc Type","values":["text"]},{"key":"thesis:degree_discipline","label":"Discipline","values":["Biochemistry"]},{"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":["Chemistry, Biochemistry","Biophysics, General"]}]},{"id":"language_rights","label":"Language and Rights","entries":[{"key":"dc:language","label":"Dc Language","values":["eng"]},{"key":"dc:rights","label":"Dc Rights","values":["Copyright 1989 Stayton, Patrick Sean"]}]},{"id":"identifiers","label":"Identifiers","entries":[{"key":"dc:identifier","label":"Identifier","values":["AAI9011038","(UMI)AAI9011038","http://hdl.handle.net/2142/20509"]}]},{"id":"additional","label":"Additional Metadata","entries":[{"key":"dc:description","label":"Description","values":["The cytochrome P-450$\\sb{\\rm cam}$ reaction cycle is a complex set of coordinated chemical transformations requiring precise temporal and spatial control of reactivities. Cytochrome P-450$\\sb{\\rm cam}$ catalyzes the regio- and stereo-specific hydroxylation of camphor to form 5-exo-hydroxycamphor. The two reducing equivalents required for this reaction are supplied physiologically by putidaredoxin, a Fe$\\sb2$S$\\sb2$ iron-sulfur protein. The mammalian cytochromes P-450 are also known to interact with cytochrome b$\\sb5$, a small redox protein for which a high resolution crystal structure is available. To characterize the molecular cytochrome P-450$\\sb{\\rm cam}$ binding surface, cytochrome b$\\sb5$ was first genetically engineered to afford a fluorescent derivative capable of monitoring its association with cytochrome P-450$\\sb{\\rm cam}$. The interaction was subsequently computer modeled by looking for van der Waals complementarity and salt bridge formation between the cytochrome b$\\sb5$ anionic binding surface and basic residues on the cytochrome P-450$\\sb{\\rm cam}$ surface. A good fit was found on the proximal surface of nearest approach to the cytochrome P-450$\\sb{\\rm cam}$ heme prosthetic group.","Subsequent binding competition studies demonstrated that putidaredoxin competitively inhibits the cytochrome b$\\sb5$-cytochrome P-450$\\sb{\\rm cam}$ association, suggesting that the same P-450$\\sb{\\rm cam}$ surface is utilized by both partners. Site directed mutagenesis of the modeled basic residues suggested that this is indeed the site of putidaredoxin-cytochrome P-450$\\sb{\\rm cam}$ association. Further time resolved fluorescence studies on the putidaredoxin C-terminal tryptophan suggested that this residue is located near an anionic protein surface. This data supports a complex model featuring electrostatic complementarity between an anionic putidaredoxin surface and the cationic cytochrome P-450$\\sb{\\rm cam}$ binding surface, with the essential tryptophan positioned to mediate electron transfer.","Made available in DSpace on 2011-05-07T12:41:15Z (GMT). No. of bitstreams: 2 license.txt: 4922 bytes, checksum: 910b249b4beec47e7ab768910c8f966f (MD5) 9011038.pdf: 6223770 bytes, checksum: 4c96a7806b0245ca093b1ac5c0bb667b (MD5) Previous issue date: 1989","Item marked as restricted to the 'UIUC Users [automated]' Group (id=2) by Howard Ding (hding2@illinois.edu) on 2011-05-07T14:44:22Z Item is restricted indefinitely.","Restriction data tranferred 2014-07-01T11:19:32-05:00 Original Data Group with Access UIUC Users [automated] Release Date: none Reason: ETDs are only available to UIUC Users without author permission","ETDs are only available to UIUC Users without author permission","U of I Only"]},{"key":"dc:title","label":"Title","values":["Macromolecular recognition in the cytochrome P450(cam) enzyme system"]}]}],"canonical_facts":{"dc:contributor":["Sligar, Stephen G."],"dc:creator":["Stayton, Patrick Sean"],"dc:date":["2011-05-07T12:41:15Z","10000-01-01","1989"],"dc:description":["The cytochrome P-450$\\sb{\\rm cam}$ reaction cycle is a complex set of coordinated chemical transformations requiring precise temporal and spatial control of reactivities. Cytochrome P-450$\\sb{\\rm cam}$ catalyzes the regio- and stereo-specific hydroxylation of camphor to form 5-exo-hydroxycamphor. The two reducing equivalents required for this reaction are supplied physiologically by putidaredoxin, a Fe$\\sb2$S$\\sb2$ iron-sulfur protein. The mammalian cytochromes P-450 are also known to interact with cytochrome b$\\sb5$, a small redox protein for which a high resolution crystal structure is available. To characterize the molecular cytochrome P-450$\\sb{\\rm cam}$ binding surface, cytochrome b$\\sb5$ was first genetically engineered to afford a fluorescent derivative capable of monitoring its association with cytochrome P-450$\\sb{\\rm cam}$. The interaction was subsequently computer modeled by looking for van der Waals complementarity and salt bridge formation between the cytochrome b$\\sb5$ anionic binding surface and basic residues on the cytochrome P-450$\\sb{\\rm cam}$ surface. A good fit was found on the proximal surface of nearest approach to the cytochrome P-450$\\sb{\\rm cam}$ heme prosthetic group.","Subsequent binding competition studies demonstrated that putidaredoxin competitively inhibits the cytochrome b$\\sb5$-cytochrome P-450$\\sb{\\rm cam}$ association, suggesting that the same P-450$\\sb{\\rm cam}$ surface is utilized by both partners. Site directed mutagenesis of the modeled basic residues suggested that this is indeed the site of putidaredoxin-cytochrome P-450$\\sb{\\rm cam}$ association. Further time resolved fluorescence studies on the putidaredoxin C-terminal tryptophan suggested that this residue is located near an anionic protein surface. This data supports a complex model featuring electrostatic complementarity between an anionic putidaredoxin surface and the cationic cytochrome P-450$\\sb{\\rm cam}$ binding surface, with the essential tryptophan positioned to mediate electron transfer.","Made available in DSpace on 2011-05-07T12:41:15Z (GMT). No. of bitstreams: 2 license.txt: 4922 bytes, checksum: 910b249b4beec47e7ab768910c8f966f (MD5) 9011038.pdf: 6223770 bytes, checksum: 4c96a7806b0245ca093b1ac5c0bb667b (MD5) Previous issue date: 1989","Item marked as restricted to the 'UIUC Users [automated]' Group (id=2) by Howard Ding (hding2@illinois.edu) on 2011-05-07T14:44:22Z Item is restricted indefinitely.","Restriction data tranferred 2014-07-01T11:19:32-05:00 Original Data Group with Access UIUC Users [automated] Release Date: none Reason: ETDs are only available to UIUC Users without author permission","ETDs are only available to UIUC Users without author permission","U of I Only"],"dc:identifier":["AAI9011038","(UMI)AAI9011038","http://hdl.handle.net/2142/20509"],"dc:language":["eng"],"dc:rights":["Copyright 1989 Stayton, Patrick Sean"],"dc:subject":["Chemistry, Biochemistry","Biophysics, General"],"dc:title":["Macromolecular recognition in the cytochrome P450(cam) enzyme system"],"dc:type":["text"],"thesis:degree_discipline":["Biochemistry"],"thesis:degree_level":["Dissertation"],"thesis:degree_name":["Ph.D."],"thesis:institution_name":["University of Illinois at Urbana-Champaign"]},"updated_at":"2026-07-22T22:25:16Z"}