{"id":{"repo_id":"uiuc","oai_identifier":"oai:www.ideals.illinois.edu:2142/72355"},"canonical_url":"https://search.dev.ndltd.org/etd/uiuc/oai:www.ideals.illinois.edu:2142/72355","repository":{"repo_id":"uiuc","name":"University of Illinois - Urbana-Champaign","base_url":"https://www.ideals.illinois.edu/oai-pmh"},"display":{"title":"Thelin Operon in Pseudomonas Incognita: Cloning, Nucleotide Sequence and Deduced Amino Acid Sequence of Cytochrome P-450lin and Other Enzymes Involved in the Catabolism of Linalool","abstract":"Cytochrome P-450 catalyzed reactions are extremely important in the metabolic pathways of both pro- and eucaryotes. Detailed mechanistic understanding of this superfamily, however, has been hampered by the availability of only one well characterized procaryotic system. Preliminary characterization of cytochrome P-450lin (P-450lin), the enzyme responsible for the 8-methyl hydroxylation of linalool as the first committed step of Pseudomonas incognita's utilization of that substrate as its sole carbon source, indicates its importance in expanding the foundation of the P-450 superfamily. Paramount to detailed mechanistic dissection, however, is the availability of a genetic handle, reagent quantities of pure enzyme and a reliable tertiary structure. Herein, we describe the realization of the first two goals which should aid in the completion of the third. Utilizing a Polymerase Chain Reaction-based cloning strategy based on the P-450lin NH$\\sb2$-terminal and tryptic-fragment amino acid sequence, the cytochrome P-450lin operon (lin) was cloned. Four overlapping clones representing 10.8 kb of novel sequence were isolated and sequenced. Eight open reading frames were found in this sequence. The GenBank, EMBL, and SwissProt databases were searched in order to identify the translated sequences. Their identities based on the database search are as follows: (1) inconclusive, (2) hydratase, (3) P-450lin, (4) regulatory, (5) linalool-8-aldehyde dehydrogenase, (6) linalool-8-alcohol dehydrogenase, (7) lin-redoxin and (8) lin-redoxin reductase. The identities of the P-450lin, lin-redoxin, lin-redoxin reductase and linalool-8-alcohol dehydrogenase were also confirmed by the presence of the anticipated NH$\\sb2$-terminal amino acid sequence determined for these proteins previously. A putative promoter sequence has been tentatively identified preceding OPF3. An optimized amino acid sequence alignment of P-450lin with cytochrome P-450cam shows the two enzymes to have only 25% identity. Based on the low sequence identity between P-450lin and all other P-450 sequences ($<$30%), P-450lin has been identified as the sole member of a new prokaryotic P-450 family, CYP111, in accordance with the nomenclature guidelines for the P-450 superfamily.","abstract_html":"Cytochrome P-450 catalyzed reactions are extremely important in the metabolic pathways of both pro- and eucaryotes. Detailed mechanistic understanding of this superfamily, however, has been hampered by the availability of only one well characterized procaryotic system. Preliminary characterization of cytochrome P-450lin (P-450lin), the enzyme responsible for the 8-methyl hydroxylation of linalool as the first committed step of Pseudomonas incognita's utilization of that substrate as its sole carbon source, indicates its importance in expanding the foundation of the P-450 superfamily. Paramount to detailed mechanistic dissection, however, is the availability of a genetic handle, reagent quantities of pure enzyme and a reliable tertiary structure. Herein, we describe the realization of the first two goals which should aid in the completion of the third. Utilizing a Polymerase Chain Reaction-based cloning strategy based on the P-450lin NH$\\sb2$-terminal and tryptic-fragment amino acid sequence, the cytochrome P-450lin operon (lin) was cloned. Four overlapping clones representing 10.8 kb of novel sequence were isolated and sequenced. Eight open reading frames were found in this sequence. The GenBank, EMBL, and SwissProt databases were searched in order to identify the translated sequences. Their identities based on the database search are as follows: (1) inconclusive, (2) hydratase, (3) P-450lin, (4) regulatory, (5) linalool-8-aldehyde dehydrogenase, (6) linalool-8-alcohol dehydrogenase, (7) lin-redoxin and (8) lin-redoxin reductase. The identities of the P-450lin, lin-redoxin, lin-redoxin reductase and linalool-8-alcohol dehydrogenase were also confirmed by the presence of the anticipated NH$\\sb2$-terminal amino acid sequence determined for these proteins previously. A putative promoter sequence has been tentatively identified preceding OPF3. An optimized amino acid sequence alignment of P-450lin with cytochrome P-450cam shows the two enzymes to have only 25% identity. Based on the low sequence identity between P-450lin and all other P-450 sequences ($<$30%), P-450lin has been identified as the sole member of a new prokaryotic P-450 family, CYP111, in accordance with the nomenclature guidelines for the P-450 superfamily.","abstract_has_math":true,"creators":["Ropp, Jerry Dezz"],"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":2014,"date_issued":"2014-12-17T21:58:42Z","date_published":"2014-12-17T21:58:42Z","updated_at":"2026-07-22T22:26:06Z","subjects":["Biology, Microbiology","Chemistry, Biochemistry"],"languages":[],"rights":[],"rights_urls":[],"identifier_entries":[{"key":"dc:identifier","label":"Identifier","values":["(UMI)AAI9314933"],"render_values":[{"text":"(UMI)AAI9314933","href":null,"code":true}]}]},"links":{"outbound_url":"http://hdl.handle.net/2142/72355","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":["Ropp, Jerry Dezz"]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"dc:date","label":"Dc Date","values":["2014-12-17T21:58:42Z","10000-01-01","1993"]},{"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":["Biology, Microbiology","Chemistry, Biochemistry"]}]},{"id":"identifiers","label":"Identifiers","entries":[{"key":"dc:identifier","label":"Identifier","values":["http://hdl.handle.net/2142/72355","(UMI)AAI9314933"]}]},{"id":"additional","label":"Additional Metadata","entries":[{"key":"dc:description","label":"Description","values":["Cytochrome P-450 catalyzed reactions are extremely important in the metabolic pathways of both pro- and eucaryotes. Detailed mechanistic understanding of this superfamily, however, has been hampered by the availability of only one well characterized procaryotic system. Preliminary characterization of cytochrome P-450lin (P-450lin), the enzyme responsible for the 8-methyl hydroxylation of linalool as the first committed step of Pseudomonas incognita's utilization of that substrate as its sole carbon source, indicates its importance in expanding the foundation of the P-450 superfamily. Paramount to detailed mechanistic dissection, however, is the availability of a genetic handle, reagent quantities of pure enzyme and a reliable tertiary structure. Herein, we describe the realization of the first two goals which should aid in the completion of the third. Utilizing a Polymerase Chain Reaction-based cloning strategy based on the P-450lin NH$\\sb2$-terminal and tryptic-fragment amino acid sequence, the cytochrome P-450lin operon (lin) was cloned. Four overlapping clones representing 10.8 kb of novel sequence were isolated and sequenced. Eight open reading frames were found in this sequence. The GenBank, EMBL, and SwissProt databases were searched in order to identify the translated sequences. Their identities based on the database search are as follows: (1) inconclusive, (2) hydratase, (3) P-450lin, (4) regulatory, (5) linalool-8-aldehyde dehydrogenase, (6) linalool-8-alcohol dehydrogenase, (7) lin-redoxin and (8) lin-redoxin reductase. The identities of the P-450lin, lin-redoxin, lin-redoxin reductase and linalool-8-alcohol dehydrogenase were also confirmed by the presence of the anticipated NH$\\sb2$-terminal amino acid sequence determined for these proteins previously. A putative promoter sequence has been tentatively identified preceding OPF3. An optimized amino acid sequence alignment of P-450lin with cytochrome P-450cam shows the two enzymes to have only 25% identity. Based on the low sequence identity between P-450lin and all other P-450 sequences ($<$30%), P-450lin has been identified as the sole member of a new prokaryotic P-450 family, CYP111, in accordance with the nomenclature guidelines for the P-450 superfamily.","Made available in DSpace on 2014-12-17T21:58:42Z (GMT). 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Detailed mechanistic understanding of this superfamily, however, has been hampered by the availability of only one well characterized procaryotic system. Preliminary characterization of cytochrome P-450lin (P-450lin), the enzyme responsible for the 8-methyl hydroxylation of linalool as the first committed step of Pseudomonas incognita's utilization of that substrate as its sole carbon source, indicates its importance in expanding the foundation of the P-450 superfamily. Paramount to detailed mechanistic dissection, however, is the availability of a genetic handle, reagent quantities of pure enzyme and a reliable tertiary structure. Herein, we describe the realization of the first two goals which should aid in the completion of the third. Utilizing a Polymerase Chain Reaction-based cloning strategy based on the P-450lin NH$\\sb2$-terminal and tryptic-fragment amino acid sequence, the cytochrome P-450lin operon (lin) was cloned. Four overlapping clones representing 10.8 kb of novel sequence were isolated and sequenced. Eight open reading frames were found in this sequence. The GenBank, EMBL, and SwissProt databases were searched in order to identify the translated sequences. Their identities based on the database search are as follows: (1) inconclusive, (2) hydratase, (3) P-450lin, (4) regulatory, (5) linalool-8-aldehyde dehydrogenase, (6) linalool-8-alcohol dehydrogenase, (7) lin-redoxin and (8) lin-redoxin reductase. The identities of the P-450lin, lin-redoxin, lin-redoxin reductase and linalool-8-alcohol dehydrogenase were also confirmed by the presence of the anticipated NH$\\sb2$-terminal amino acid sequence determined for these proteins previously. A putative promoter sequence has been tentatively identified preceding OPF3. An optimized amino acid sequence alignment of P-450lin with cytochrome P-450cam shows the two enzymes to have only 25% identity. Based on the low sequence identity between P-450lin and all other P-450 sequences ($<$30%), P-450lin has been identified as the sole member of a new prokaryotic P-450 family, CYP111, in accordance with the nomenclature guidelines for the P-450 superfamily.","Made available in DSpace on 2014-12-17T21:58:42Z (GMT). No. of bitstreams: 1 9314933.pdf: 7603707 bytes, checksum: 95cfe5ed922f8b7125f9a5d713064f7b (MD5) Previous issue date: 1993","Embargo set by: Seth Robbins for item 72523 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","215 p.","Thesis (Ph.D.)--University of Illinois at Urbana-Champaign, 1993."],"dc:identifier":["http://hdl.handle.net/2142/72355","(UMI)AAI9314933"],"dc:subject":["Biology, Microbiology","Chemistry, Biochemistry"],"dc:title":["Thelin Operon in Pseudomonas Incognita: Cloning, Nucleotide Sequence and Deduced Amino Acid Sequence of Cytochrome P-450lin and Other Enzymes Involved in the Catabolism of Linalool"],"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":"2026-07-22T22:26:06Z"}