Purification and characterization of the alternative nitrogenase from the photosynthetic bacterium Rhodospirillum rubrum.
The alternative nitrogenase from a nifH mutant of the photosynthetic bacterium Rhodospirillum rubrum has been purified and characterized. The dinitrogenase protein (ANF1) contains three subunits in an apparent alpha2beta2gamma2 structure and contains Fe but no Mo or V. A factor capable of activating apo-dinitrogenase (lacking the FeMo cofactor) from Azotobacter vinelandii was extracted from the alternative dinitrogenase protein with N-methylformamide. The electron paramagnetic resonance (EPR) signal of the dinitrogenase protein is not characteristic of the EPR signals of molybdenum- or vanadium-containing dinitrogenases. The alternative dinitrogenase reductase (ANF2) was purified as an alpha2 dimer containing an Fe4S4 cluster and exhibited an EPR spectrum characteristic of dinitrogenase reductases. The enzyme complex reduces protons to H2 very well but reduces N2 to ammonium poorly. Acetylene is reduced to a mixture of ethylene and ethane.
Davis
R
R
Department of Biochemistry, University of Wisconsin, Madison 53706, USA.
Lehman
L
L
Petrovich
R
R
Shah
V K
VK
Roberts
G P
GP
Ludden
P W
PW
D000114:Acetylene / Q000378:metabolism; D001051:Apoenzymes / Q000737:chemistry / Q000378:metabolism; D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism; D017037:Dinitrogenase Reductase / Q000737:chemistry / Q000378:metabolism; D004578:Electron Spin Resonance Spectroscopy; D004591:Electrophoresis, Polyacrylamide Gel; D004980:Ethane / Q000378:metabolism; D007501:Iron / Q000032:analysis; D007506:Iron-Sulfur Proteins / Q000737:chemistry* / Q000378:metabolism; D007527:Isoenzymes / Q000737:chemistry* / Q000378:metabolism; D008670:Metals / Q000032:analysis; D009154:Mutation; D009591:Nitrogenase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D010084:Oxidation-Reduction; D010088:Oxidoreductases; D011487:Protein Conformation; D000644:Quaternary Ammonium Compounds / Q000378:metabolism; D012247:Rhodospirillum rubrum / Q000201:enzymology* / Q000235:genetics; D013379:Substrate Specificity; D014233:Tricarboxylic Acids / Q000032:analysis
D001051:Apoenzymes; D001426:Bacterial Proteins; D017037:Dinitrogenase Reductase; D007506:Iron-Sulfur Proteins; D007527:Isoenzymes; D008670:Metals; D000644:Quaternary Ammonium Compounds; D014233:Tricarboxylic Acids; C028143:homocitric acid; D007501:Iron; D010088:Oxidoreductases; D009591:Nitrogenase; C029130:nitrogenase reductase; D004980:Ethane; D000114:Acetylene