Electroneutral Na+-2 Cl−-Leucine Cotransport by Lobster Hepatopancreatic Brush-Border Membrane Vesicles

Abstract

Uptake of L-[3H]leucine by lobster hepatopancreatic brush-border membrane vesicles was stimulated by a transmembrane NaCl gradient (o>i), but not by identical gradients of NaSCN or other Cl− salts (e.g. K+, Li+, NH4+, Cs+ or choline), suggesting that amino acid transfer depended upon both Na+ and Cl−. In NaCl medium at acidic pH, leucine uptake was largely electroneutral and unresponsive to a transmembrane potential generated by permeable anions; however, in Na+-free medium, amino acid transport was strongly electrogenic under the same conditions. Leucine influx occurred by a combination of two carrier processes at physiologically acidic pH. One exhibited an influx Kt of 0.59 mmol l−1, a JM of 390 pmol mg protein−1 S−1 and a cotransport stoichiometry of 1 Na+: 2 Cl+: 1 leucine. This process was most strongly cis-inhibited by the non-polar amino acids phenylalanine, methionine and isoleucine, and most weakly inhibited by the more polar species methylaminoisobutyric acid (MeAIB), hydroxyproline, glutamate and arginine. The second leucine carrier system showed a very low binding affinity and could not be distinguished from diffusion, was Na+- and Cl−-independent, and was cis-inhibited by more polar amino acids such as lysine, hydroxyproline, MeAIB, alanine and glutamate. These results suggest that brush-border leucine transport in lobster hepatopancreas at acidic pH may occur by a combination of a modified L-system, that includes ion cosubstrates, and either by a second undefined Na+-independent process with a broad structural specificity or by multiple Na+-independent processes.