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

Abstract

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 1−1, a JM of 390pmol 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 nonpolar amino acids phenylalanine, methionine and isoleucine, and most weakly inhibited by the more polar species methylaminoisobutyric acid (MeAlB), 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.