{"id":{"repo_id":"uiuc","oai_identifier":"oai:www.ideals.illinois.edu:2142/70522"},"canonical_url":"https://search.dev.ndltd.org/etd/uiuc/oai:www.ideals.illinois.edu:2142/70522","repository":{"repo_id":"uiuc","name":"University of Illinois - Urbana-Champaign","base_url":"https://www.ideals.illinois.edu/oai-pmh"},"display":{"title":"Active Center Structure and Sequence Studies on Bacterial Luciferase Utilizing the Essential Cysteine, Protease-Labile Region, and Delta Fragment","abstract":"This study examines the structure of the active center of bacterial luciferase by exploiting the existence of two well-documented structural features of the (alpha) subunit of the enzyme, the essential cysteine and the protease-labile region, which are thought to reside there. Chemical modification of this cysteinyl residue causes slight perturbations, dependent upon the structure of the modifying reagent, in the structure of the protease-labile region as well as loss of enzymatic activity, as does intrasubunit crosslinking induced by the reagent p-azidophenacyl bromide. High concentrations of phosphate ion or the reaction product FMN slightly decrease the reactivity of the essential cysteine. These results and those of limited proteolysis of luciferase modified at this cysteine with ('3)H-N-ethylmaleimide, N-terminal sequence analysis of the resulting fragments, and specific cleavage of (alpha) at the essential thiol caused by modification with 2-nitro-5-thiocyanatobenzoic acid, are consistent with a model which places this residue on an exposed polypeptide loop, which is a portion of the active center, (TURN)100 residues from the amino terminus of (alpha). The protease-labile region appears to be composed of two sections of primary structure, one located on the same polypeptide loop as the essential cysteine (the "slow" area), and the other (TURN)100-130 residues from the carboxy terminus (the "fast" area). Amino acid sequences of peptides derived from the proteolytic fragments of the "(delta)" family are also presented.","abstract_html":"This study examines the structure of the active center of bacterial luciferase by exploiting the existence of two well-documented structural features of the (alpha) subunit of the enzyme, the essential cysteine and the protease-labile region, which are thought to reside there. Chemical modification of this cysteinyl residue causes slight perturbations, dependent upon the structure of the modifying reagent, in the structure of the protease-labile region as well as loss of enzymatic activity, as does intrasubunit crosslinking induced by the reagent p-azidophenacyl bromide. High concentrations of phosphate ion or the reaction product FMN slightly decrease the reactivity of the essential cysteine. These results and those of limited proteolysis of luciferase modified at this cysteine with ('3)H-N-ethylmaleimide, N-terminal sequence analysis of the resulting fragments, and specific cleavage of (alpha) at the essential thiol caused by modification with 2-nitro-5-thiocyanatobenzoic acid, are consistent with a model which places this residue on an exposed polypeptide loop, which is a portion of the active center, (TURN)100 residues from the amino terminus of (alpha). The protease-labile region appears to be composed of two sections of primary structure, one located on the same polypeptide loop as the essential cysteine (the "slow" area), and the other (TURN)100-130 residues from the carboxy terminus (the "fast" area). Amino acid sequences of peptides derived from the proteolytic fragments of the "(delta)" family are also presented.","abstract_has_math":false,"creators":["Rausch, Steven Kirk"],"institution":"University of Illinois at Urbana-Champaign","degree_name":"Ph.D.","degree_level":"Dissertation","degree_discipline":"Biochemistry","degree_department":null,"school":null,"contributors":[],"advisors":[],"committee_chairs":[],"committee_members":[],"year":2014,"date_issued":"2014-12-15T23:43:40Z","date_published":"2014-12-15T23:43:40Z","updated_at":"2026-07-22T22:26:03Z","subjects":["Chemistry, Biochemistry"],"languages":[],"rights":[],"rights_urls":[],"identifier_entries":[{"key":"dc:identifier","label":"Identifier","values":["(UMI)AAI8310002"],"render_values":[{"text":"(UMI)AAI8310002","href":null,"code":true}]}]},"links":{"outbound_url":"http://hdl.handle.net/2142/70522","outbound_label":"Handle","outbound_source":"dc:identifier"},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:creator","label":"Author","values":["Rausch, Steven Kirk"]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"dc:date","label":"Dc Date","values":["2014-12-15T23:43:40Z","10000-01-01","1983"]},{"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"]}]},{"id":"identifiers","label":"Identifiers","entries":[{"key":"dc:identifier","label":"Identifier","values":["http://hdl.handle.net/2142/70522","(UMI)AAI8310002"]}]},{"id":"additional","label":"Additional Metadata","entries":[{"key":"dc:description","label":"Description","values":["This study examines the structure of the active center of bacterial luciferase by exploiting the existence of two well-documented structural features of the (alpha) subunit of the enzyme, the essential cysteine and the protease-labile region, which are thought to reside there. Chemical modification of this cysteinyl residue causes slight perturbations, dependent upon the structure of the modifying reagent, in the structure of the protease-labile region as well as loss of enzymatic activity, as does intrasubunit crosslinking induced by the reagent p-azidophenacyl bromide. High concentrations of phosphate ion or the reaction product FMN slightly decrease the reactivity of the essential cysteine. These results and those of limited proteolysis of luciferase modified at this cysteine with ('3)H-N-ethylmaleimide, N-terminal sequence analysis of the resulting fragments, and specific cleavage of (alpha) at the essential thiol caused by modification with 2-nitro-5-thiocyanatobenzoic acid, are consistent with a model which places this residue on an exposed polypeptide loop, which is a portion of the active center, (TURN)100 residues from the amino terminus of (alpha). The protease-labile region appears to be composed of two sections of primary structure, one located on the same polypeptide loop as the essential cysteine (the "slow" area), and the other (TURN)100-130 residues from the carboxy terminus (the "fast" area). Amino acid sequences of peptides derived from the proteolytic fragments of the "(delta)" family are also presented.","Made available in DSpace on 2014-12-15T23:43:40Z (GMT). No. of bitstreams: 1 8310002.pdf: 6963189 bytes, checksum: 71104157a0f67774074679fdbab5b011 (MD5) Previous issue date: 1983","Embargo set by: Seth Robbins for item 70688 Lift date: Forever Reason: Restricted to the U of I community idenfinitely during batch ingest of legacy ETDs","Restricted to the U of I community idenfinitely during batch ingest of legacy ETDs","U of I Only","238 p.","Thesis (Ph.D.)--University of Illinois at Urbana-Champaign, 1983."]},{"key":"dc:title","label":"Title","values":["Active Center Structure and Sequence Studies on Bacterial Luciferase Utilizing the Essential Cysteine, Protease-Labile Region, and Delta Fragment"]}]}],"canonical_facts":{"dc:creator":["Rausch, Steven Kirk"],"dc:date":["2014-12-15T23:43:40Z","10000-01-01","1983"],"dc:description":["This study examines the structure of the active center of bacterial luciferase by exploiting the existence of two well-documented structural features of the (alpha) subunit of the enzyme, the essential cysteine and the protease-labile region, which are thought to reside there. Chemical modification of this cysteinyl residue causes slight perturbations, dependent upon the structure of the modifying reagent, in the structure of the protease-labile region as well as loss of enzymatic activity, as does intrasubunit crosslinking induced by the reagent p-azidophenacyl bromide. High concentrations of phosphate ion or the reaction product FMN slightly decrease the reactivity of the essential cysteine. These results and those of limited proteolysis of luciferase modified at this cysteine with ('3)H-N-ethylmaleimide, N-terminal sequence analysis of the resulting fragments, and specific cleavage of (alpha) at the essential thiol caused by modification with 2-nitro-5-thiocyanatobenzoic acid, are consistent with a model which places this residue on an exposed polypeptide loop, which is a portion of the active center, (TURN)100 residues from the amino terminus of (alpha). The protease-labile region appears to be composed of two sections of primary structure, one located on the same polypeptide loop as the essential cysteine (the "slow" area), and the other (TURN)100-130 residues from the carboxy terminus (the "fast" area). Amino acid sequences of peptides derived from the proteolytic fragments of the "(delta)" family are also presented.","Made available in DSpace on 2014-12-15T23:43:40Z (GMT). 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