Multiple transport pathways for dibasic amino acids in the larval midgut of the silkworm Bombyx mori

The transport pathways for dibasic amino acids were investigated in brush b order membrane vesicles (BBMV) from the anterior-middle (AM) and posterior (P) regions of Bombyx mori midgut. In the absence of K+, a low-affinity sat urable transport of arginine in both AM- and P-BBMV (K-m 1.01 mM, V-max 4.0 7 nmol/7s/mg protein and K-m 1.38 mM, V-max 2.26 nmol/7s/mg protein, respec tively) was detected. Arginine influx was dependent on the membrane electri cal potential (Delta psi) and increased raising the alkalinity of the exter nal medium from pH 7.2 to 10.6. Competition experiments indicated the follo wing order of substrate affinity: arginine, homoarginine, N-G-monomethylarg inine, N-G-nitroarginine > lysine much greater than omithine > cysteine > m ethionine, Leucine, valine and BCH (2-amino-2-norbornanecarboxylic acid) di d not inhibit arginine influx. In the presence of external K+. the influx o f arginine as a function of arginine concentration fitted to a complex satu ration kinetics compatible with both a low-affinity and a high-affinity com ponent. The latter (K-m 0.035 mM, V-max 2.54 nmol/7s/mg protein) was fully characterized. The influx rate had an optimum at pH 8.8, was strongly affec ted by Delta psi and was homogeneous along the midgut. The substrate affini ty rank was: homoarginine > arginine, N-G-monomethylarginine much greater t han cysteine, lysine much greater thanN(G)-nitroarginine > ornithine > meth ionine. Leucine and amino acids with a hydrophobic side chain were not acce pted. This system is also operative in the absence of potassium, with the s ame order of specificity but a very low activity. Lysine influx is mediated by two more transport systems, the leucine uniport and the K+/leucine symp ort specific for amino acids with a hydrophobic side chain that recognizes lysine at extravesicular pH values (pH(out)) exceeding 9. Both the uniport and the symport differ from the cationic transport systems so far identifie d in mammals because they are unaffected by N-ethylmaleimide, have no signi ficant affinity for neutral amino acids in the presence of the cation and s how a striking difference in their optimum pH. (C) 2001 Elsevier Science Lt d. All rights reserved.