VFe nitrogenase

Recently, a second or alternative nitrogenase has been isolated from Azotobacter vinelandii (21) and Azotobacter chroococcum (22) that contains vanadium as opposed to molybdenum. The MoFe and VFe nitrogenase proteins from A. vinelandii (called Av and. 4vl , respectively) are known to have different polypeptide structures and it obviously of interest to know to what extent the cluster composition is conserved. Variable temperature MCD studies of the as isolated and thionine oxidized proteins provided a convenient means of addressing this question. 

Finally, the FeMoeo is transferred to nifDK (apo-MoFe protein) to form the holo-MoFe protein with the help of ATP and several proteins nifH, 7, and GroEL. The X-ray crystal structure of nifDK shows a positively charged channel through which FeMoeo could enter. " The mf system for synthesis of the FeVco is thought to be similar to the nif system, with added assistance in the last step by vnfG, a third subunit of VFe nitrogenase that is necessary for activity. 

NifM is required for maturation of VnfH, and NifS and U seem to be important for provision of sulfide and probably iron for the biosynthesis of the vanadium nitrogenase. The apo VFe protein has been isolated from an A. vinelandii strain deleted for nifKD and nifB (169). It was an hexamer that could be activated in vitro by the addi-... 

The work described above was greatly aided by the provision of samples from colleagues D. gigas Ni(II)-substituted rubredoxin, I. Moura and J. J. G. Moura T. thermophilus ferredoxin, J. A. Fee VFe and MoFe nitrogenase proteins from A. vinelandii, B. J. Hales. Research in the author s laboratory is supported by grants from NIH (GM33806) and NSF (DMB8796212) and an Alfred P. Sloan Research Fellowship. 

Nitrogenase, MoFe-, VFe-, and Fe-only classes Burgess (1984), Orme-Johnson (1985)... 

The only kinetic study on a V nitrogenase [74] reports that the rates of binding of the reduced Fe protein with the VFe protein and the subsequent electron-transfer rates are very similar to those for Mo nitrogenase, as the components of the V nitrogenase form a slightly weaker electron-transfer complex. 

The vanadium nitrogenase has been isolated from A. vinelandii and A. chroococcum (Table 1). With the molybdenum in the protein replaced by vanadium, the vanadium nitrogenase resembles its molybdenum counterpart. It has the molybdenum-iron (MoFe) protein equivalent in the vanadium-iron (VFe) protein, as well as the iron (Fe) protein equivalent. 

The presence of an open reading frame between vnfD and vnfK led to the identification of an additional small subunit type (8, encoded by vnfG) of the VFe protein it has the hexameric structure 012 2 2 ill). A homologous gene exists between anfD and anfK (13a) and encodes a small subunit of the third nitrogenase (R. Eady and R. Pau, unpublished). 

Fig, 1. Schematic representation of the a and /3 polypeptides of the MoFe protein, the VFe protein, and the third nitrogenase. The vertical lines represent the position of the five invariant Cys residues of the a subunit and the three invariant Cys residues of the /3 subunit. The flanking amino acid sequences are shown to indicate the degree of sequence homology around these Cys residues. The wide horizontal lines represent the polypeptide chains of the subunits of the three nitrogenases (see Ref 25). 

MoFe protein in K. pneumoniae VFe proteins Third nitrogenase of A. vinelandii ... 

The components of the third nitrogenase, analogous to the MoFe and VFe proteins (dinitrogenase-3) have been purified and partially characterized (5). The third nitrogenase can be isolated in two active forms with different subunit configurations, ai/ 2 and a2(32 there is a 8 subunit present at uncertain stoichiometry encoded by anfG (R. Eady and R. Pau, unpublished). 

Scheme 2. MgATP-dependent electron transfer between components of V nitrogenase, The pre-steady-state electron transfer reactions between the Fe protein and the VFe protein of V nitrogenase of A, chroococcum have been analyzed in terms of this scheme (52). Ac2 represents the Fe protein and Acl represents the VFe protein of this system. This scheme is analogous to that used in the detailed study of Mo nitrogenase (see Hef. 50).

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