Cloning, nucleotide sequencing, and expression of the Azospirillum brasilense lon gene: involvement in iron uptake

Abstract

The lon gene of Escherichia coli encodes the lon (La) protease, which is associated with cellular protein degradation. A lon gene homolog from Azospirillum brasilense, a nitrogen-fixing soil bacterium which lives in association with the roots of cereal grasses, was cloned and characterized. The nucleotide sequence of the A. brasilense lon gene was determined. It contains an open reading frame that encodes a protein of 810 amino acids with a predicted molecular mass of about 90 kDa. The deduced amino acid sequence showed a high level of homology with the sequences of all the known lon gene products. An open reading frame homologous to the E. coli clpX gene was found in front of the lon gene. Transcriptional analysis showed that the lon gene of A. brasilense is induced by heat shock and that the mRNA is monocistronic. An A. brasilense mutant, with Tn5 inserted in the lon gene, was shown to be defective in iron uptake and failed to express two membrane proteins that are induced by iron starvation in the parental strain.