THE BKDR GENE OF PSEUDOMONAS-PUTIDA IS REQUIRED FOR EXPRESSION OF THEBKD 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 Ps eudomonas putida. The structural genes encoding all four proteins of t he bkd operon have been cloned, and their nucleotide sequences have be en determined (G. Burns, K. T. Madhusudhan, K. Hatter, and J. R. Sokat ch, p. 177-184 in S. Silver, A. M. Chakrabarty, B. Iglewski, and S. Ka plan [ed.], Pseudomonas: Biotransformations, Pathogenesis, and Evolvin g Biotechnology, American Society for Microbiology, Washington D.C., 1 990). An open reading frame which encoded a protein with 36.5% amino a cid identity to the leucine-responsive regulatory protein (Lrp) of Esc herichia coli was found immediately upstream of the bkd operon. Chromo somal mutations affecting this gene, named bkdR, resulted in a loss of ability to use branched-chain amino acids as carbon and energy source s and failure to produce branched-chain keto acid dehydrogenase. These mutations were complemented in trans by plasmids which contained inta ct 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 m ust be positive. Mutations affecting bkdR could also be complemented b y plasmids containing lrp of E. coli. This is the first instance of a Lrp-like protein demonstrated to regulate expression of an operon outs ide of E. coli.