The nitrogenase structural genes (nifHDK) of the endophytic diazotroph Herbaspirillum seropedicae were isolated from a genomic bank by plate hybridization. Sequence analysis of the DNA showed a consensus promoter region upstream for the nifH gene containing a -24/-12 type promoter together with NifA- and integration host factor (IHF)- binding sites. The derived protein sequences of NifH, NifD and NifK contained conserved cysteine residues for binding iron-sulfur clusters and the iron-molybdenum cofactor. These protein sequences showed the strongest similarities to the nifHDK gene products of the symbiotic diazotroph Bradyrhizobium japonicum (93.5%, 91.3% and 83.3%, respectively), the plant-associated diazotroph Azospirillum brasilense (90.0%, 83.7% and 75.1%, respectively) and to Thiobacillus ferrooxidans (91.0%, 83.4% and 81.1%, respectively) of the same phylogenetic group of the protobacteria.