The ntrBC genes of Azospirillum brasilense are part of a nifR3-like-ntrB-ntrC operon and are negatively regulated.
A cosmid able to complement the Nif- and nitrate-dependent growth phenotypes of the Azospirillum brasilense mutant FP9 was isolated from a genomic library of the wild-type strain FP2. A 6-kb DNA region was sequenced and showed two open reading frames (ORFs) identified as the ntrB and ntrC genes. An ORF1 located upstream from the ntrB gene and coding for a 36-kDa polypeptide showed similarity to the nifR3 gene of Rhodobacter capsulatus and the ORF1 of Rhizobium leguminosarum, both located upstream from the ntrB gene in a complex operon. Two other unidentified ORFs (ORF5 and partial ORF4) coding for hydrophobic polypeptides were also observed. delta ORF1-ntrBC, ORF1, ntrB, and ntrC mutants obtained by recombination of suicide plasmids containing an insertion of a promoterless lacZ kanamycin cassette showed decreased nitrogenase activities and were unable to grow on nitrate as the sole N source. These phenotypes were restored by complementation with plasmids containing the ntrC gene. Analysis of lacZ transcriptional fusions suggested that the ORF1-ntrBC operon in Azospirillum brasilense is expressed from a promoter located upstream from the ORF1 and that it is negatively regulated by the ntrC gene product.
Machado
H B
HB
Department of Pharmacology, Universidade Federal do Paraná, Curitiba, PR, Brazil.
Yates
M G
MG
Funayama
S
S
Rigo
L U
LU
Steffens
M B
MB
Souza
E M
EM
Pedrosa
F O
FO
D000595:Amino Acid Sequence; D015806:Azospirillum brasilense / Q000235:genetics*; D001483:Base Sequence; D002874:Chromosome Mapping; D003001:Cloning, Molecular; D003360:Cosmids; D004269:DNA, Bacterial / Q000235:genetics; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D005816:Genetic Complementation Test; D007763:Lac Operon; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009586:Nitrogen Fixation / Q000235:genetics; D016366:Open Reading Frames; D009876:Operon; D011401:Promoter Regions, Genetic; D017386:Sequence Homology, Amino Acid
D004269:DNA, Bacterial