L-leucine, L-methionine, and L-phenylalanine share a Na+/K+-dependent amino acid transporter in shrimp hepatopancreas

摘要

Hepatopancreatic brush border membrane vesicles (BBMV), made from Atlantic White shrimp (Litopenaeus setiferus), were used to characterize the transport properties of H-3-l-leucine influx by these membrane systems and how other essential amino acids and the cations, sodium and potassium, interact with this transport system. H-3-l-leucine uptake by BBMV was pH-sensitive and occurred against transient transmembrane concentration gradients in both Na+- and K+-containing incubation media, suggesting that either cation was capable of providing a driving force for amino acid accumulation. H-3-l-leucine uptake in NaCl or KCl media were each three times greater in acidic pH (pH 5.5) than in alkaline pH (pH 8.5). The essential amino acid, l-methionine, at 20 mM significantly (p < 0.0001) inhibited the 2-min uptakes of 1 mM H-3-l-leucine in both Na+- and K+-containing incubation media. The residual H-3-l-leucine uptake in the two media were significantly greater than zero (p < 0.001), but not significantly different from each other (p > 0.05) and may represent an l-methionine- and cation-independent transport system. H-3-l-leucine influxes in both NaCl and KCl incubation media were hyperbolic functions of [l-leucine], following the carrier-mediated Michaelis-Menten equation. In NaCl, H-3-l-leucine influx displayed a low apparent K (M) (high affinity) and low apparent J (max), while in KCl the transport exhibited a high apparent K (M) (low affinity) and high apparent J (max). l-methionine or l-phenylalanine (7 and 20 mM) were competitive inhibitors of H-3-l-leucine influxes in both NaCl and KCl media, producing a significant (p < 0.01) increase in H-3-l-leucine influx K (M), but no significant response in H-3-l-leucine influx J (max). Potassium was a competitive inhibitor of sodium co-transport with H-3-l-leucine, significantly (p < 0.01) increasing H-3-l-leucine influx K (M) in the presence of sodium, but having negligible effect on H-3-l-leucine influx J (max) in the same medium. These results suggest that shrimp BBMV transport H-3-l-leucine by a single l-methionine- and l-phenylalanine-shared carrier system that is enhanced by acidic pH and can be stimulated by either Na+ or K+ acting as co-transport drivers binding to shared activator sites.