IDENTIFICATION OF A NEW TRANSPORT-SYSTEM (Y+L) IN HUMAN ERYTHROCYTES THAT RECOGNIZES LYSINE AND LEUCINE WITH HIGH-AFFINITY

1. The effect of neutral amino acids on the transport Of L-lysine across the human erythrocyte membrane was studied. 2. All neutral amino acids tested (range 0.3-5 mM) inhibit the influx of L-[C-14]lysine (1-mu-M). The inhibition pattern is biphasic, and tends to reach a maximum at approximately 50 % of the original flux. The concentrations that give 25 % inhibition are (mM): L-cysteine (2.7), L-alanine (1.3), L-serine (0.9), L-isoleucine (0.6), L-phenylalanine (0.35), L-methionine (< 0.3), L-leucine (< 0.3). L-lysine and L-arginine completely inhibit the rate at the highest concentration. 3. These results can be explained by assuming that L-lysine transport occurs through two independent transporters that differ in their affinity for neutral amino acids. A detailed kinetic analysis of the effect Of L-leucine on L-lysine entry is consistent with this hypothesis. 4. Using a new experimental strategy, the substrate and inhibitor transport parameters for the two systems were determined. The half-saturation constants for lysine (+/- S.E.M.) are found to be: K(mA), 0.014 +/- 0.002 mM and K(mB), 0.112 +/- 0.017 mM. The maximum rates differ by a factor of 8.2 (V(maxB)/V(maxA)). The leucine inhibition constants are: K(iA), 0.022 +/- 0.003 mm and K(iB), 30.36 +/- 7.9 mm. If the sodium in the incubation medium is replaced by potassium, the apparent affinity for leucine (1/K(iA)) is reduced approximately 30-fold. 5. The maximum inhibition caused by leucine decreases as the lysine concentration is raised, showing that leucine acts upon the higher affinity system. 6. When added to the trans side, L-leucine, L-phenylalanine and L-isoleucine do not cause inhibition, but stimulate the flux by approximately 30 %. This indicates that these analogues are also transported. 7. In conclusion, in the concentration range 1-100-mu-M, lysine crosses the red cell membrane through two distinct transport systems, one of which recognizes both neutral and cationic amino acids with high affinity.