Klebsiella pneumoniae nitrogenases

Thomley, R.N.F. and Lowe, D.J. (1984) The mechanism of Klebsiella pneumonia nitrogenase action. Determination of rate constants required for the simulation of the kinetic ofNj reduction and H2 evolution, Biochem. J. 224, 895-901. 

Mo K-edge EXAFS spectrum (left panel) and EXAFS Fourier transform (right panel) of Klebsiella pneumoniae nitrogenase MoFe protein. The solid line is the processed experimental spectrum and the dashed line a calculated one. ... 

Kala-Azar, 278 Keggin anions, 1034 electrochemistry, 1050 Keggin structures, 1035 heteropolyanions, 1037 Klebsiella pneumoniae nitrogenases, 1425 Kroll process, 325 Krypton compounds difluorides, 313 fluoride, 313... 

The nitrogenase proteins are generally characterized by two letters indicating the species and strains of bacteria and the numerals 1 for the MoFe protein and 2 for the Fe protein. Thus, the Fe protein from Azotobacter vinelandii is Av2 and the MoFe protein from Klebsiella pneumoniae is Kpl. 

G Lihong, M. Madden,VK. Shah, RH. Burris (1990) Citrate substitutes for homocitrate in nitrogenase of a nifV mutant of Klebsiella pneumoniae. Biochemistry, 29 8577-8581... 

Mayer, S.M., Lawson, D.M., Gormal, C.A., Roe, S.M. and Smith B.E. (1999) New insights into structure-function relationships in nitrogenase A 1.6 A resolution X-ray crystallographic study of Klebsiella pneumoniae MoFe-protein, J. Mol Biol., 292, 871-891. 

Not only are two molecules of ATP hydrolyzed to pump each electron, but the Fe-protein must receive electrons from a powerful (low E°) reductant such as reduced ferredoxin, reduced flavodoxin, or dithionite. Klebsiella pneumoniae contains a pyruvate flavodoxin oxidoreductase (Eq. 15-35) that reduces either flavodoxin or ferredoxin to provide the low potential electron donor.29 30 In some bacteria, e.g., the strictly aerobic Azotobacter, NADPH is the electron donor for reduction of N2. The Fe-protein is thought to accept electrons from a chain that includes at least the ordinary bacterial ferredoxin (Fd) and a special one-electron-accepting azotoflavin, a flavoprotein that is somewhat larger than the flavodoxins (Chapter 15) and appears to play a specific role in N2 fixation.31 In Clostridium and Rhizobium reduced ferredoxins generated by cleavage of pyruvate reduce nitrogenase directly.32... 

Figure 24-4 Sequence of nif genes of Klebsiella pneumoniae.56 These precede the his operon directly at the right side. The nitrogenase structural genes are marked with green.

The principle advances in the area has been made using x-ray structural analysis. Crystallographic data have been first produced for the nitrogenase complex of FeP (A2) and FeMoP (Al) from Azotobacter vinelandii (Kim and Rees, 1992) and for the corresponding complex of Cp2 and Cpl from Clostridium pasterianum (Bolen et al., 1993), ). A 1.6 A resolution X-ray crystallographic structure of Klebsiella pneumoniae proteins has been recently reported (Mayer et al., 1999) It was shown that FeMoco sites in Al, Cpl, and Kpl are 70 A apart and FeMoco and P clusters are separated by about 19 A. X-ray structures of the nitrogenase complex and the active site clusters are presented in (Figs. 3.2-3.4). 

Thomeley, R.N.F., Ashy, G., Howarth, J.V., Millar, N.C., and Gutfreund, H. (1989) A transient kinetic study of the nitrogenase of Klebsiella pneumonia by stopped-flow calorimetry. Comparison with the myosin ATPase, Biochem. J. 264, 657-661. 

Smith, B. E. Studies on the iron-molybdenum cofactor from the nitrogenase Mo-Fe protein of Klebsiella pneumoniae, in Ref. 7, p. 179... 

Pre-steady-state stopped-flow and rapid quench techniques applied to Mo nitrogenase have provided powerful approaches to the study of this complex enzyme. These studies of Klebsiella pneumoniae Mo nitrogenase showed that a pre-steady-state burst in ATP hydrolysis accompanied electron transfer from the Fe protein to the MoFe protein, and that during the reduction of N2 an enzyme-bound dinitrogen hydride was formed, which under denaturing conditions could be trapped as hydrazine. A comprehensive model developed from a computer simulation of the kinetics of these reactions and the kinetics of the pre-steady-state rates of product formation (H2, NH3) led to the formulation of Scheme 1, the Thorneley and Lowe scheme (50) for nitrogenase function. 

Fig. 2. The fe -weighted EXAFS data associated with the iron K-edge of the iron-molybdenum cofactor (FeMoco) extracted from the FeMo-protein of the nitrogenase of Klebsiella pneumoniae, and its Fourier transform (19). 

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