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 border membrane vesicles (BBMV) from the anterior-middle (AM) and posterior (P) regions of Bombyx mori midgut. In the absence of K(+), a low-affinity saturable transport of arginine in both AM- and P-BBMV (K(m) 1.01 mM, V(max) 4.07 nmol/7s/mg protein and K(m) 1.38 mM, V(max) 2.26 nmol/7s/mg protein, respectively) was detected. Arginine influx was dependent on the membrane electrical potential (Deltapsi) and increased raising the alkalinity of the external medium from pH 7.2 to 10.6. Competition experiments indicated the following order of substrate affinity: arginine, homoarginine, N(G)-monomethylarginine, N(G)-nitroarginine>lysine>>ornithine>cysteine>methionine. Leucine, valine and BCH (2-amino-2-norbornanecarboxylic acid) did not inhibit arginine influx. In the presence of external K(+), the influx of arginine as a function of arginine concentration fitted to a complex saturation kinetics compatible with both a low-affinity and a high-affinity component. 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 affected by Deltapsi and was homogeneous along the midgut. The substrate affinity rank was: homoarginine>arginine, N(G)-monomethylarginine>>cysteine, lysine>>N(G)-nitroarginine>ornithine>methionine. Leucine and amino acids with a hydrophobic side chain were not accepted. This system is also operative in the absence of potassium, with the same order of specificity but a very low activity. Lysine influx is mediated by two more transport systems, the leucine uniport and the K(+)/leucine symport 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 identified in mammals because they are unaffected by N-ethylmaleimide, have no significant affinity for neutral amino acids in the presence of the cation and show a striking difference in their optimum pH.
Department of Biology, University of Milan, via Celoria 26, 20133 Milan, Italy.
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Casartelli
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Leonardi
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Fiandra
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Parenti
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Giordana
D000468:Alkalies / Q000378:metabolism; D000599:Amino Acids, Diamino / Q000378:metabolism*; D000818:Animals; D001120:Arginine / Q000378:metabolism; D001665:Binding Sites; D001692:Biological Transport; D012831:Bombyx / Q000378:metabolism*; D002414:Cations, Monovalent; D004064:Digestive System / Q000378:metabolism; D005033:Ethylmaleimide / Q000494:pharmacology; D007814:Larva; D008239:Lysine / Q000378:metabolism; D011188:Potassium / Q000378:metabolism
D000468:Alkalies; D000599:Amino Acids, Diamino; D002414:Cations, Monovalent; D001120:Arginine; D008239:Lysine; D005033:Ethylmaleimide; D011188:Potassium