STRUCTURAL AND FUNCTIONAL-ANALYSIS OF THE FIXLJ GENES OF RHIZOBIUM-LEGUMINOSARUM BIOVAR PHASEOLI CNPAF512

The fixLJ genes of Rhizobium leguminosarum biovar phaseoli CNPAF512 we re identified by DNA hybridization of a genomic library with an intern al fragment of the Rhizobium Meliloti fixJ gene. The nucleotide sequen ce was determined and the corresponding amino acid sequence was aligne d with the amino acid sequences of the FixL proteins of R, meliloti, B radyrhizobium japonicum and Azorhizobium caulinodans. While the FixJ p rotein and the carboxy-terminal part of the FixL protein are highly ho mologous to the other FixL and FixJ proteins, the homology in the cent ral heme-binding, oxygen-sensing domain and in the amino-terminal doma in of FixL is very low. The R. leguminosarum bv. phaseoli FixL protein does not contain the heme-binding motif defined for the previously de scribed FixL proteins. R. leguminosarum bv. phaseoli fixLJ and fixJ mu tants were constructed. These mutants can still fix nitrogen, albeit a t a reduced level. Expression analysis of nifA-gusA and nifH-gusA fusi ons in the constructed mutants revealed that the R. leguminosarum bv. phaseoli fixLJ genes are involved in microaerobic nifH expression but not in nifA expression.