Substrate specificity of the brush border K+-leucine symport of Bombyx mori larval midgut.
L-leucine uptake into membrane vesicles from Bombyx mori larval midgut was tested for inhibition by 55 compounds, which included sugars, N-methylated, alpha-, beta-, gamma-, delta-, epsilon-amino acids, primary amines, alpha-amino alcohols, monocarboxylic organic acids and alpha-ketoacids. Based on cis-inhibition experiments performed at the high pH (10.8) characteristic of the midgut luminal content in vivo, we find that the carrier binding site interacts with molecules which possess a well-defined set of structural features. Amino acids are preferentially accepted as anions and the ideal inhibitor must have an hydrophobic region and a polar head constituted by a chiral carbon atom bearing two hydrophilic groups, a deprotonated amino-group and a dissociated carboxylic group. Binding is reduced if one of the two hydrophilic groups is removed. Lowering the pH to less alkaline value (8.8) only affects the affinity of delta- and epsilon-amino acids, which are excluded from binding because of their positively charged side-chain. Modifications of the potassium electrochemical gradient increased the affinity constant values of the molecules, but have little effect on the rank of specificity. Physiological implications of the data reported are discussed.
Parenti
P
P
Department of Environmental Sciences, University of Milano-Bicocca, Milan, Italy. paolo.parenti@unimi.it
Forcella
M
M
Pugliese
A
A
Casartelli
M
M
Giordana
B
B
Leonardi
M G
MG
Hanozet
G M
GM
D026905:Amino Acid Transport Systems; D026923:Amino Acid Transport Systems, Basic; D026921:Amino Acid Transport Systems, Neutral; D000818:Animals; D012831:Bombyx / Q000378:metabolism*; D002352:Carrier Proteins / Q000037:antagonists & inhibitors* / Q000378:metabolism; D004064:Digestive System; D004563:Electrochemistry; D006863:Hydrogen-Ion Concentration; D007814:Larva; D007930:Leucine / Q000378:metabolism*; D008871:Microvilli / Q000378:metabolism; D011188:Potassium / Q000378:metabolism*; D013379:Substrate Specificity
D026905:Amino Acid Transport Systems; D026923:Amino Acid Transport Systems, Basic; D026921:Amino Acid Transport Systems, Neutral; D002352:Carrier Proteins; D007930:Leucine; D011188:Potassium