Genetic separation of high- and low-affinity transport systems for branched-chain amino acids in Escherichia coli K-12

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Abstract
The E. coli K-12 mutant strain AE4107 (livH::Mu) is defective in the high-affinity binding protein-mediated uptake system for L-leucine, L-valine and L-isoleucine (LIV-I). This strain was used to produce mutations in the residual LIV-II membrane-bound branched-chain amino acid uptake system. Mutants selected for their inability to utilize exogenous L-leucine were defective in the LIV-II system and fell into 2 classes. One class, represented by strain AE410709 (livP9), showed a complete loss of saturable uptake for L-leucine, L-valine and L-isoleucine up to 50 .mu.M, and a 2nd class, represented by strain AE4017012 (liv-12), showed a residual component of saturable leucine uptake with increased Km. These mutations, livP9 and liv-12, were closely linked and mapped in the 74-78 min region of the E. coli genetic map. Strains constructed so that they lacked both LIV-I and LIV-II transport systems excreted leucine. Strains of the genotype livH+ livP had normal high-affinity binding protein-mediated transport (LIV-I and leucine specific), but the low-affinity (LIV-II) transport was completely missing. The high-affinity binding protein-mediated transport systems (LIV-I and leucine specific) can apparently operate independently of the membrane-bound LIV-II system.