Genetic analysis of the Pseudomonas aeruginosa PAO high-affinity branched-chain amino acid transport system by use of plasmids carrying the bra genes

Abstract

About 30 mutants of Pseudomonas aeruginosa PAO defective in the high-affinity branched-chain amino acid transport system (LIV-I) were isolated by the selection for resistance to 4-aza-DL-leucine, a toxic leucine analog for LIV-I. All of the mutants were complemented by plasmid pKTH24, harboring the braC gene, which encodes the branched-chain amino acid-binding protein, and the four open reading frames named braD, braE, braF, and braG (T. Hoshino and K. Kose, J. Bacteriol. 172:5531-5539, 1990). We identified five cistrons corresponding to these bra genes by complementation analysis with various derivatives of pKTH24, confirming that the braD, braE, braF, and braG genes are required for the LIV-I transport system. We also found mutations that seem likely to be mutations in a promoter region for the bra genes and those with polarity in the intercistronic region between braC and braD. Analysis with an omega interposon showed that the bra genes are organized as an operon and are cotranscribed in the order braC-braD-braE-braF-braG from a promoter located in the 5'-flanking region of the braC gene.