Repression and inhibition of transport systems for branched-chain amino acids in Salmonella typhimurium

Abstract

Kinetics of the transport systems common for entry of L-isoleucine, L-leucine and L-valine in S. typhimurium LT2 were analyzed as a function of substrate concentration in the range of 0.5-45 .mu.M. The systems of transport mutants KA203 (ilvT3) and KA204 (ilvT4) are composed of 2 components; apparent Km values for uptake of isoleucine, leucine and valine by the low Km component are 2 .mu.M, 2-3 .mu.M and 1 .mu.M, respectively, and by the high Km component 30 .mu.M, 20-40 .mu.M and 0.1 mM, respectively. The transport system(s) of the wild type was not separated into components but displays single Km values of 9 .mu.M for isoleucine, 10 .mu.M for leucine and 30 .mu.M for valine. The transport activity of the wild type was repressed by L-leucine, .alpha.-ketoisocaproate, glycyl-L-isoleucine, glycyl-L-leucine and glycyl-L-methionine. That for the transport mutants was repressed by L-alanine, L-isoleucine, L-methionine, L-valine, .alpha.-ketoisovalerate, .alpha.-keto-.beta.-methylvalerate, glycyl-L-alanine, glycyl-L-threonine and glycyl-L-valine, besides the compounds described above. Repression of the mutant transport systems resulted in disappearance of the low Km component for valine uptake, together with a decrease in Vmax of the high Km component; the kinetic analysis with isoleucine and leucine as substrates was not possible because of poor uptake. The maximum reduction of the transport activity for isoleucine was obtained after growing cells for 2-3 generations in a medium supplemented with repressor, and for the derepression, protein synthesis was essential after removal of the repressor. The transport activity for labeled isoleucine in the transport mutant and wild-type strains was inhibited by unlabeled L-alanine, L-cysteine, L-isoleucine, L-leucine, L-methionine, L-threonine and L-valine. D-Amino acids did not repress or inhibit the transport activity of cells for entry of isoleucine.