Molecular cloning and sequencing of an operon, carRS of Azospirillum brasilense, that codes for a novel two-component regulatory system: demonstration of a positive regulatory role of carR for global control of carbohydrate catabolism.
A pleiotropic carbohydrate mutant, CR17, of Azospirillum brasilense RG (wild type) that assimilates C4 dicarboxylates (succinate and malate) but not carbohydrate (fructose, arabinose, galactose, glycerol, and gluconate) as C sources for growth was used to identify the car (carbohydrate regulation) locus by complementation analysis. The 2.8-kb genomic fragment that complemented the Car- defect of CR17 and overlapped the fru operon (S. Chattopadhyay, A. Mukherjee, and S. Ghosh, J. Bacteriol. 175:3240-3243, 1993) has now been completely sequenced. The sequence contains an operon, carRS, coding for two proteins, CARR and CARS, having 236 and 352 amino acid residues, respectively. The 3'-flanking region of the carRS operon showed sequence homology with the 5' terminus of the fruB gene of a related bacterium, Rhodobacter capsulatus. A complementation study with carRS deletion clones showed that only the carR+ gene was required to complement the Car- defect of CR17, signifying that the carbohydrate pleiotropy was due to a lesion within this gene. Although the 2.8-kb DNA containing the carRS operon when introduced by conjugation into CR17 also complemented the Car- defect, the complemented transconjugant was unable to utilize succinate as a C source. The reason for this is not clear. A sequence analysis of the two protein products strongly suggests that the protein pair may constitute a novel two-component regulatory system for global expression of carbohydrate catabolic pathways in A. brasilense.
Chattopadhyay
S
S
Department of Biochemistry, Bose Institute, Calcutta, India.
Mukherjee
A
A
Ghosh
S
S
D000595:Amino Acid Sequence; D015806:Azospirillum brasilense / Q000235:genetics* / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics*; D001483:Base Sequence; D050260:Carbohydrate Metabolism; D002352:Carrier Proteins / Q000235:genetics; D015964:Gene Expression Regulation, Bacterial; D005816:Genetic Complementation Test; D008040:Genetic Linkage; D047908:Intracellular Signaling Peptides and Proteins; D008969:Molecular Sequence Data; D009876:Operon / Q000235:genetics*; D015183:Restriction Mapping; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D014157:Transcription Factors
D001426:Bacterial Proteins; C091096:CarR protein, Azospirillum brasilense; C091094:CarS protein, Myxococcus xanthus; D002352:Carrier Proteins; D047908:Intracellular Signaling Peptides and Proteins; D014157:Transcription Factors; C028640:fructose-induced HPr-like protein, bacteria