THE GENES ENCODING THE DELTA SUBUNITS OF DINITROGENASE-2 AND DINITROGENASE-3 ARE REQUIRED FOR MO-INDEPENDENT DIAZOTROPHIC GROWTH BY AZOTOBACTER-VINELANDII

vnfG and anfG encode the delta subunits of alternative nitrogenases 2 and 3 in Azotobacter vinelandii, respectively. As a first step towards elucidating the role of these subunits, diazotrophic growth and acetylene reduction studies were conducted on mutants containing alterations in the genes encoding these subunits. Mutants containing a stop codon (C36stop) or an in-frame deletion in anfG were unable to grow in N-free, Mo-deficient medium (Anf(-)). Mutants in which cysteine 36 of AnfG (a residue conserved between VnfG and AnfG) was changed to Ala or Ser were Anf(+). Thus, this conserved cysteine is not essential for the function of AnfG in dinitrogenase 3. A mutant with a stop codon in vnfG (C17stop) grew after a lag of 25 h in N-free, Mo-deficient medium containing V2O5. However, a Nif(-) Anf(-) strain with this mutation was unable to grow under these conditions. This shows that the vnfG gene product is required for nitrogenase 2-dependent growth. Strains with mutations in vnfG and anfG reduced acetylene to different degrees. This indicates that the delta subunits are not required for acetylene reduction by nitrogenases 2 and 3.