Identification of a new transport system (y+L) in human erythrocytes that recognizes lysine and leucine with high affinity.

Abstract

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‐[14C]lysine (1 microM). 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: KmA, 0.014 +/‐ 0.002 mM and KmB, 0.112 +/‐ 0.017 mM. The maximum rates differ by a factor of 8.2 (VmaxB/VmaxA). The leucine inhibition constants are: KiA, 0.022 +/‐ 0.003 mM and KiB, 30.36 +/‐ 7.9 mM. If the sodium in the incubation medium is replaced by potassium, the apparent affinity for leucine (1/KiA) 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 microM, lysine crosses the red cell membrane through two distinct transport systems, one of which recognizes both neutral and cationic amino acids with high affinity.