{"id":{"repo_id":"vt","oai_identifier":"oai:vtechworks.lib.vt.edu:10919/36177"},"canonical_url":"https://search.dev.ndltd.org/etd/vt/oai:vtechworks.lib.vt.edu:10919/36177","repository":{"repo_id":"vt","name":"Virginia Tech","base_url":"https://vtechworks.lib.vt.edu/oai/request"},"display":{"title":"Characterization of an altered MoFe protein from a nifV- strain from Azotobacter vinelandii","abstract":"The site of substrate binding and reduction for the nitrogenase complex is located on the iron molybdenum cofactor (FeMo-co) which is contained within the a-subunit of the molybdenum iron protein. FeMo co consists of a metal sulfur core composed of an FeS cluster bridged by three inorganic sulfides to a MoFeS cluster. An organic acid, homocitrate, is coordinated to the Mo atom through its 2-carboxy and 2-hydroxy groups. Homocitrate is formed by the condensation of acetyl-CoA and a-ketoglutarate, which is catalyzed by a homocitrate synthase encoded by nifV. By deleting the nifV gene from Azotobacter vinelandii we were able to study the role of homocitrate in nitrogenase catalysis. A poly-histidine tail was incorporated into the C-termini of the a-subunit permitting isolation of the homocitrateless MoFe protein by using metal affinity chromatography. We have found that the addition of a poly-histidine tag does not alter the catalytic behavior of the native enzyme. In NifV- strains of Klebsiella pneumoniae, citrate has been found to replace homocitrate as the organic constituent of FeMo-co. We have found no evidence this is so in A. vinelandii. Gas chromatography mass spectrophotometry studies indicate little or no organic acids are associated with FeMo-co. We examined the catalytic properties of the NifV- MoFe protein In the mutant, H2 evolution is inhibited by the addition of CO, unlike in the wild type. We have found that the NifV- MoFe protein from A. vinelandii is able to catalyze the reduction of acetylene to both ethylene and ethane.","abstract_html":"The site of substrate binding and reduction for the nitrogenase complex is located on the iron molybdenum cofactor (FeMo-co) which is contained within the a-subunit of the molybdenum iron protein. FeMo co consists of a metal sulfur core composed of an FeS cluster bridged by three inorganic sulfides to a MoFeS cluster. An organic acid, homocitrate, is coordinated to the Mo atom through its 2-carboxy and 2-hydroxy groups. Homocitrate is formed by the condensation of acetyl-CoA and a-ketoglutarate, which is catalyzed by a homocitrate synthase encoded by nifV. By deleting the nifV gene from Azotobacter vinelandii we were able to study the role of homocitrate in nitrogenase catalysis. A poly-histidine tail was incorporated into the C-termini of the a-subunit permitting isolation of the homocitrateless MoFe protein by using metal affinity chromatography. We have found that the addition of a poly-histidine tag does not alter the catalytic behavior of the native enzyme. In NifV- strains of Klebsiella pneumoniae, citrate has been found to replace homocitrate as the organic constituent of FeMo-co. We have found no evidence this is so in A. vinelandii. Gas chromatography mass spectrophotometry studies indicate little or no organic acids are associated with FeMo-co. We examined the catalytic properties of the NifV- MoFe protein In the mutant, H2 evolution is inhibited by the addition of CO, unlike in the wild type. We have found that the NifV- MoFe protein from A. vinelandii is able to catalyze the reduction of acetylene to both ethylene and ethane.","abstract_has_math":false,"creators":["Comaratta, Leonard M."],"institution":"Virginia Tech","degree_name":"Master of Science","degree_level":"masters","degree_discipline":"Biochemistry and Anaerobic Microbiology","degree_department":"Biochemistry and Anaerobic Microbiology","school":null,"contributors":[],"advisors":[],"committee_chairs":["Dean, Dennis R."],"committee_members":["Larson, Timothy J.","Gregory, Eugene M.","Chen, Jiann-Shin","Claus, George William"],"year":1998,"date_issued":"1998-12-03","date_published":"1998-12-03","updated_at":"2026-07-22T22:19:18Z","subjects":["MoFe","Azotobacter vinelandii","Nitrogenase"],"languages":[],"rights":["In Copyright"],"rights_urls":["http://rightsstatements.org/vocab/InC/1.0/"],"identifier_entries":[{"key":"dc:identifier.other","label":"Dc Identifier Other","values":["etd-121598-113534"],"render_values":[{"text":"etd-121598-113534","href":null,"code":true}]}]},"links":{"outbound_url":"http://hdl.handle.net/10919/36177","outbound_label":"Handle","outbound_source":"dc:identifier.uri"},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:contributor.committeechair","label":"Committee Chair","values":["Dean, Dennis R."]},{"key":"dc:contributor.committeemember","label":"Committee Member","values":["Larson, Timothy J.","Gregory, Eugene M.","Chen, Jiann-Shin","Claus, George William"]},{"key":"dc:contributor.department","label":"Department","values":["Biochemistry and Anaerobic Microbiology"]},{"key":"dc:creator","label":"Author","values":["Comaratta, Leonard M."]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"dc:date.accessioned","label":"Dc Date Accessioned","values":["2014-03-14T20:50:01Z"]},{"key":"dc:date.available","label":"Dc Date Available","values":["2014-03-14T20:50:01Z","1999-11-13"]},{"key":"dc:date.issued","label":"Date","values":["1998-12-03"]},{"key":"dc:publisher","label":"Institution","values":["Virginia Tech"]},{"key":"dc:type","label":"Dc Type","values":["Thesis"]},{"key":"thesis:degree_discipline","label":"Discipline","values":["Biochemistry and Anaerobic Microbiology"]},{"key":"thesis:degree_level","label":"Degree Level","values":["masters"]},{"key":"thesis:degree_name","label":"Degree Name","values":["Master of Science"]},{"key":"thesis:institution_name","label":"Thesis Institution Name","values":["Virginia Polytechnic Institute and State University"]}]},{"id":"subjects_keywords","label":"Subjects and Keywords","entries":[{"key":"dc:subject","label":"Dc Subject","values":["MoFe","Azotobacter vinelandii","Nitrogenase"]}]},{"id":"language_rights","label":"Language and Rights","entries":[{"key":"dc:rights","label":"Dc Rights","values":["In Copyright"]},{"key":"dc:rights.uri","label":"Rights URI","values":["http://rightsstatements.org/vocab/InC/1.0/"]}]},{"id":"identifiers","label":"Identifiers","entries":[{"key":"dc:identifier.other","label":"Dc Identifier Other","values":["etd-121598-113534"]},{"key":"dc:identifier.uri","label":"Identifier URI","values":["http://hdl.handle.net/10919/36177"]}]},{"id":"additional","label":"Additional Metadata","entries":[{"key":"dc:description.abstract","label":"Abstract","values":["The site of substrate binding and reduction for the nitrogenase complex is located on the iron molybdenum cofactor (FeMo-co) which is contained within the a-subunit of the molybdenum iron protein. FeMo co consists of a metal sulfur core composed of an FeS cluster bridged by three inorganic sulfides to a MoFeS cluster. An organic acid, homocitrate, is coordinated to the Mo atom through its 2-carboxy and 2-hydroxy groups. Homocitrate is formed by the condensation of acetyl-CoA and a-ketoglutarate, which is catalyzed by a homocitrate synthase encoded by nifV. By deleting the nifV gene from Azotobacter vinelandii we were able to study the role of homocitrate in nitrogenase catalysis. A poly-histidine tail was incorporated into the C-termini of the a-subunit permitting isolation of the homocitrateless MoFe protein by using metal affinity chromatography. We have found that the addition of a poly-histidine tag does not alter the catalytic behavior of the native enzyme. In NifV- strains of Klebsiella pneumoniae, citrate has been found to replace homocitrate as the organic constituent of FeMo-co. We have found no evidence this is so in A. vinelandii. Gas chromatography mass spectrophotometry studies indicate little or no organic acids are associated with FeMo-co. We examined the catalytic properties of the NifV- MoFe protein In the mutant, H2 evolution is inhibited by the addition of CO, unlike in the wild type. We have found that the NifV- MoFe protein from A. vinelandii is able to catalyze the reduction of acetylene to both ethylene and ethane."]},{"key":"dc:description.degree","label":"Dc Description Degree","values":["Master of Science"]},{"key":"dc:title","label":"Title","values":["Characterization of an altered MoFe protein from a nifV- strain from Azotobacter vinelandii"]}]}],"canonical_facts":{"dc:contributor.committeechair":["Dean, Dennis R."],"dc:contributor.committeemember":["Larson, Timothy J.","Gregory, Eugene M.","Chen, Jiann-Shin","Claus, George William"],"dc:contributor.department":["Biochemistry and Anaerobic Microbiology"],"dc:creator":["Comaratta, Leonard M."],"dc:date.accessioned":["2014-03-14T20:50:01Z"],"dc:date.available":["2014-03-14T20:50:01Z","1999-11-13"],"dc:date.issued":["1998-12-03"],"dc:description.abstract":["The site of substrate binding and reduction for the nitrogenase complex is located on the iron molybdenum cofactor (FeMo-co) which is contained within the a-subunit of the molybdenum iron protein. FeMo co consists of a metal sulfur core composed of an FeS cluster bridged by three inorganic sulfides to a MoFeS cluster. An organic acid, homocitrate, is coordinated to the Mo atom through its 2-carboxy and 2-hydroxy groups. Homocitrate is formed by the condensation of acetyl-CoA and a-ketoglutarate, which is catalyzed by a homocitrate synthase encoded by nifV. By deleting the nifV gene from Azotobacter vinelandii we were able to study the role of homocitrate in nitrogenase catalysis. A poly-histidine tail was incorporated into the C-termini of the a-subunit permitting isolation of the homocitrateless MoFe protein by using metal affinity chromatography. We have found that the addition of a poly-histidine tag does not alter the catalytic behavior of the native enzyme. In NifV- strains of Klebsiella pneumoniae, citrate has been found to replace homocitrate as the organic constituent of FeMo-co. We have found no evidence this is so in A. vinelandii. Gas chromatography mass spectrophotometry studies indicate little or no organic acids are associated with FeMo-co. We examined the catalytic properties of the NifV- MoFe protein In the mutant, H2 evolution is inhibited by the addition of CO, unlike in the wild type. We have found that the NifV- MoFe protein from A. vinelandii is able to catalyze the reduction of acetylene to both ethylene and ethane."],"dc:description.degree":["Master of Science"],"dc:identifier.other":["etd-121598-113534"],"dc:identifier.uri":["http://hdl.handle.net/10919/36177"],"dc:publisher":["Virginia Tech"],"dc:rights":["In Copyright"],"dc:rights.uri":["http://rightsstatements.org/vocab/InC/1.0/"],"dc:subject":["MoFe","Azotobacter vinelandii","Nitrogenase"],"dc:title":["Characterization of an altered MoFe protein from a nifV- strain from Azotobacter vinelandii"],"dc:type":["Thesis"],"thesis:degree_discipline":["Biochemistry and Anaerobic Microbiology"],"thesis:degree_level":["masters"],"thesis:degree_name":["Master of Science"],"thesis:institution_name":["Virginia Polytechnic Institute and State University"]},"updated_at":"2026-07-22T22:19:18Z"}