The bkdR gene of Pseudomonas putida is required for expression of the bkd operon and encodes a protein related to Lrp of Escherichia coli.
Branched-chain keto acid dehydrogenase is a multienzyme complex which is required for the metabolism of the branched-chain amino acids in Pseudomonas putida. The structural genes encoding all four proteins of the bkd operon have been cloned, and their nucleotide sequences have been determined (G. Burns, K. T. Madhusudhan, K. Hatter, and J. R. Sokatch, p. 177-184 in S. Silver, A. M. Chakrabarty, B. Iglewski, and S. Kaplan [ed.], Pseudomonas: Biotransformations, Pathogenesis, and Evolving Biotechnology, American Society for Microbiology, Washington D.C., 1990). An open reading frame which encoded a protein with 36.5% amino acid identity to the leucine-responsive regulatory protein (Lrp) of Escherichia coli was found immediately upstream of the bkd operon. Chromosomal mutations affecting this gene, named bkdR, resulted in a loss of ability to use branched-chain amino acids as carbon and energy sources and failure to produce branched-chain keto acid dehydrogenase. These mutations were complemented in trans by plasmids which contained intact bkdR. Mutations affecting bkdR did not have any effect on transport of branched-chain amino acids or transamination. Therefore, the bkdR gene product must affect expression of the bkd operon and regulation must be positive. Mutations affecting bkdR could also be complemented by plasmids containing lrp of E. coli. This is the first instance of a Lrp-like protein demonstrated to regulate expression of an operon outside of E. coli.
Madhusudhan
K T
KT
Department of Biochemistry and Molecular Biology, University of Oklahoma Health Sciences Center, Oklahoma City 73190.
Lorenz
D
D
Sokatch
J R
JR
D042942:3-Methyl-2-Oxobutanoate Dehydrogenase (Lipoamide); D000595:Amino Acid Sequence; D000597:Amino Acids, Branched-Chain / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics*; D001483:Base Sequence; D001692:Biological Transport; D004268:DNA-Binding Proteins / Q000235:genetics*; D029968:Escherichia coli Proteins; D005798:Genes, Bacterial / Q000235:genetics*; D005816:Genetic Complementation Test; D007658:Ketone Oxidoreductases / Q000235:genetics*; D051756:Leucine-Responsive Regulatory Protein; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes / Q000235:genetics*; D016366:Open Reading Frames / Q000235:genetics; D009876:Operon / Q000235:genetics; D016958:Pseudomonas putida / Q000235:genetics*; D017386:Sequence Homology, Amino Acid; D000637:Transaminases / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics*
D000597:Amino Acids, Branched-Chain; D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; C497278:Lrp protein, E coli; D009097:Multienzyme Complexes; D014157:Transcription Factors; D051756:Leucine-Responsive Regulatory Protein; D007658:Ketone Oxidoreductases; D042942:3-Methyl-2-Oxobutanoate Dehydrogenase (Lipoamide); D000637:Transaminases