Virginia Tech
Characterization of an altered MoFe protein from a nifV- strain from Azotobacter vinelandii
Abstract
dc:description.abstractThe 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.
Degree
thesis:*- Name thesis:degree_name
- Master of Science
- Level thesis:degree_level
- masters
- Discipline thesis:degree_discipline
- Biochemistry and Anaerobic Microbiology
- Department dc:contributor.department
- Biochemistry and Anaerobic Microbiology
- Grantor dc:publisher
- Virginia Tech
- Year dc:date.issued
- 1998
Author and committee
dc:creator, dc:contributor.*- Author dc:creator
-
- Comaratta, Leonard M.
- Chair dc:contributor.committeechair
-
- Dean, Dennis R.
- Committee members dc:contributor.committeemember
-
- Larson, Timothy J.
- Gregory, Eugene M.
- Chen, Jiann-Shin
- Claus, George William
Subjects
dc:subject × 3Rights
dc:rights- Statement dc:rights
-
- In Copyright
- Licence dc:rights.uri
Identifiers
dc:identifier.*- Dc Identifier Other
- etd-121598-113534
- OAI identifier oai:identifier
- oai:vtechworks.lib.vt.edu:10919/36177