Characterization of a FeMo cofactor-deficient MoFe protein from a nifE-deleted strain (DJ35) of Azotobacter vinelandii

2005 ◽  
Vol 50 (20) ◽  
pp. 2305 ◽  
Author(s):  
Ying ZHAO
Keyword(s):  
Azotobacter Vinelandii ◽  
Mofe Protein ◽  
Femo Cofactor

scholarly journals Evidence for a dynamic role for homocitrate during nitrogen fixation: the effect of substitution at the α-Lys426 position in MoFe-protein of Azotobacter vinelandii

2006 ◽  
Vol 397 (2) ◽  
pp. 261-270 ◽  
Author(s):  
Marcus C. Durrant ◽  
Amanda Francis ◽  
David J. Lowe ◽  
William E. Newton ◽  
Karl Fisher
Keyword(s):  
Active Site ◽  
Ring Opening ◽  
Azotobacter Vinelandii ◽  
Chelate Ring ◽  
Wild Type ◽  
Biologically Relevant ◽  
Mofe Protein ◽  
Key Players ◽  
Femo Cofactor ◽  
The Subject

Although it is generally accepted that the active site of nitrogenase is located on the FeMo-cofactor, the exact site(s) of N2 binding and reduction remain the subject of continuing debate, with both molybdenum and iron atoms being suggested as key players. The current consensus favours binding of acetylene and some other non-biologically relevant substrates to the central iron atoms of the FeMo-cofactor [Dos Santos, Igarashi, Lee, Hoffman, Seefeldt and Dean (2005) Acc. Chem. Res. 38, 208–214]. The reduction of N2 is, however, a more demanding process than reduction of these alternative substrates because it has a much higher activation energy and does not bind until three electrons have been accumulated on the enzyme. The possible conversion of bidentate into monodentate homocitrate on this three electron-reduced species has been proposed to free up a binding site for N2 on the molybdenum atom. One of the features of this hypothesis is that α-Lys426 facilitates chelate ring opening and subsequent orientation of the monodentate homocitrate by forming a specific hydrogen bond to the homocitrate -CH2CH2CO2− carboxylate group. In support of this concept, we show that mutation of α-Lys426 can selectively perturb N2 reduction without affecting acetylene reduction. We interpret our experimental observations in the light of a detailed molecular mechanics modelling study of the wild-type and altered MoFe-nitrogenases.

Purification and characterization of a FeMo cofactor-deficient MoFe protein

Biochemistry ◽  
1994 ◽  
Vol 33 (39) ◽  
pp. 11842-11849 ◽  
Author(s):  
Narasaiah Gavini ◽  
Li Ma ◽  
Gerald Watt ◽  
Barbara K. Burgess
Keyword(s):  
Purification And Characterization ◽  
Mofe Protein ◽  
Femo Cofactor

scholarly journals Nitrogenase MoFe protein fromClostridium pasteurianumat 1.08 Å resolution: comparison with theAzotobacter vinelandiiMoFe protein

2015 ◽  
Vol 71 (2) ◽  
pp. 274-282 ◽  
Author(s):  
Li-Mei Zhang ◽  
Christine N. Morrison ◽  
Jens T. Kaiser ◽  
Douglas C. Rees
Keyword(s):  
Active Sites ◽  
Azotobacter Vinelandii ◽  
Protein Structures ◽  
Protein Docking ◽  
Α Subunit ◽  
Mofe Protein ◽  
Fe Protein ◽  
Surrounding Environment ◽  
Solvent Environment ◽  
Femo Cofactor

The X-ray crystal structure of the nitrogenase MoFe protein fromClostridium pasteurianum(Cp1) has been determined at 1.08 Å resolution by multiwavelength anomalous diffraction phasing. Cp1 and the ortholog fromAzotobacter vinelandii(Av1) represent two distinct families of nitrogenases, differing primarily by a long insertion in the α-subunit and a deletion in the β-subunit of Cp1 relative to Av1. Comparison of these two MoFe protein structures at atomic resolution reveals conserved structural arrangements that are significant to the function of nitrogenase. The FeMo cofactors defining the active sites of the MoFe protein are essentially identical between the two proteins. The surrounding environment is also highly conserved, suggesting that this structural arrangement is crucial for nitrogen reduction. The P clusters are likewise similar, although the surrounding protein and solvent environment is less conserved relative to that of the FeMo cofactor. The P cluster and FeMo cofactor in Av1 and Cp1 are connected through a conserved water tunnel surrounded by similar secondary-structure elements. The long α-subunit insertion loop occludes the presumed Fe protein docking surface on Cp1 with few contacts to the remainder of the protein. This makes it plausible that this loop is repositioned to open up the Fe protein docking surface for complex formation.

Characterization of a FeMo cofactor-deficient MoFe protein from anifE-deleted strain (DJ35) ofAzotobacter vinelandii

2005 ◽  
Vol 50 (20) ◽  
pp. 2305-2310
Author(s):  
Ying Zhao ◽  
Shaomin Bian ◽  
Chunxi Zhang ◽  
Huina Zhou ◽  
Huangping Wang ◽  
...  
Keyword(s):  
Mofe Protein ◽  
Femo Cofactor ◽  
Ofazotobacter Vinelandii

The Immuno-Electron Microscopic Localization of the Mo-Fe Protein Component of Nitrogenase in Cells of Azotobacter Vinelandii

1973 ◽  
Vol 31 ◽  
pp. 574-575
Author(s):  
J. T. Stasny ◽  
R. C. Burns ◽  
R. W. F. Hardy
Keyword(s):  
Cellular Localization ◽  
Direct Evidence ◽  
Azotobacter Vinelandii ◽  
Intracellular Localization ◽  
Protein Component ◽  
Electron Microscopic ◽  
Thin Sections ◽  
Fe Protein ◽  
Intracytoplasmic Membranes

Structure-functlon studies of biological N2-fixation have correlated the presence of the enzyme nitrogenase with increased numbers of intracytoplasmic membranes in Azotobacter. However no direct evidence has been provided for the internal cellular localization of any nitrogenase. Recent advances concerned with the crystallizatiorTand the electron microscopic characterization of the Mo-Fe protein component of Azotobacter nitrogenase, prompted the use of this purified protein to obtain antibodies (Ab) to be conjugated to electron dense markers for the intracellular localization of the protein by electron microscopy. The present study describes the use of ferritin conjugated to goat antitMo-Fe protein immunoglobulin (IgG) and the observations following its topical application to thin sections of N2-grown Azotobacter.

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