Nitrogenase estimation

Acetylene-reduction assay Estimates nitrogenase activity by measuring the rate of acetylene reduced to ethylene. 

The process of nitrogen fixation is an essential part of the nitrogen cycle on the planet earth(l). It is estimated that greater than 60Z of the N2 that is ultimately converted to NH4+ is done so by the nitrogenase enzyme system. The availability of fixed nitrogen is often the limiting factor in plant growth( ). To... 

Figure 3-10 Estimation of the molecular mass of the polypeptide chain of the nitrogenase Fe-protein using SDS-poly-acrylamide electrophoresis from a set of four standard curves. The marker proteins are (1) catalase, (2) fumarase, (3) aldolase, (4) glyceraldehyde-phosphate dehydrogenase, (5) a-chymotrypsinogen A, and (6) myoglobin, (o) indicates position of azoferredoxin. From Nakos and Mortenson.195...

E0 = -0.042 V (Pierick et al., 1993). The reduced P-cluster of Av2 transfers at least one electron to FeMocoN to form the ESR silent "super-reduced" FeMoco, (FeMoco) 1. (Ohrme-Johnson et al., 1972 Miinck et al., 1975). A combination of cyclic voltampermetry, potentiometry and ESR spectroscopy has allowed the observation in isolated FeMoco of two redox transitions with essentially different potentials, E0 = - 0.3 eV and E0 = -1.0 eV (Newton et al., 1996). The first transition of FeMoco is from oxidize state to semireduced state of FeMoco and the second transition is related to the substrate-reducing state in FeMoco during the nitrogenase reaction turnover. Independently, for the redox pair FeMocoN ( FeMoco)2", the Eo was estimated as... 

Such cyanobacteria also have a greater need for Fe (Carr Wyman, 1986 Brand, 1991). The requirement may be intensified in nitrogen-fixers, which also need Fe as a metallic cofactor for nitrogenase. However, recent calculations (Sanudo-Wilhelmy etal., 2001) suggest that the iron requirement of nitrogenfixing organisms is much more than previously estimated. 

Several other extractants have been used to estimate plant-available Mo in soils (e.g., Cox and Kamprath, 1972 Gupta and Lipsett, 1981), but none has achieved widespread acceptance, and the ammonium oxalate extraction remains as frequently used as any other method. Biological assays have also been used to detect Mo deficiencies in soil. Franco, Peres, and Nery (1978) found that the nitrogenase activity of Azotobacter paspali was correlated with yield responses to Mo by the tropical forage legume Centrosema pubescens. 

Fixation of N2 to NH3 is catalyzed by the prokaryotic enzyme nitrogenase, which has both an electron and an ATP requirement for its functioning. Three electron pairs are required for each molecule of N2 reduced, while estimates of ATP requirements range from 2 to 20 ATP per electron utilized (Burris et al., 1980 Ljones and Burris, 1972 Bums and Hardy, 1975 Ludden and Burris, 1986). There is a measure of agreement (Schubert, 1982 Salsac et al., 1984) that 2 ATP/ probably represents a reasonable average cost in vivo. 

---