scholarly journals A role for aminopeptidase N in Na+-dependent amino acid transport in bovine renal brush-border membranes

1993 ◽  
Vol 290 (1) ◽  
pp. 59-65 ◽  
Author(s):  
S Plakidou-Dymock ◽  
M J Tanner ◽  
J D McGivan
Keyword(s):  
Amino Acid ◽  
Brush Border ◽  
Amino Acid Transport ◽  
Membrane Vesicles ◽  
Aminopeptidase N ◽  
Aminopeptidase Activity ◽  
Acid Transport ◽  
Transport Activity ◽  
Renal Brush Border Membrane ◽  
Renal Brush Border

A monoclonal antibody FD19 which removes reconstitutable Na(+)-dependent amino acid transport activity from solubilized bovine renal brush-border membrane vesicles was found to react specifically with the enzyme aminopeptidase N. Cleavage of aminopeptidase N from the membranes with papain inhibited Na(+)-dependent amino acid transport activity without affecting that of alpha-methyl D-glucoside. Removal of aminopeptidase substantially increased the Km values for the Na(+)-dependent transport of alanine, glutamine, leucine and phenylalanine without affecting the Vmax. Both Na(+)-dependent amino acid transport and aminopeptidase activity in intact vesicles were competitively inhibited by amino acids with very similar specificity. These results suggest that the amino acid-binding sites of aminopeptidase N and the transporter interact in some way to increase the Km of the transport process for its substrates. However, independent direct inactivation of the transport system by papain cannot be ruled out.

scholarly journals Reconstitution and identification of the major Na+-dependent neutral amino acid-transport protein from bovine renal brush-border membrane vesicles

1992 ◽  
Vol 281 (1) ◽  
pp. 95-102 ◽  
Author(s):  
F A Doyle ◽  
J D McGivan
Keyword(s):  
Amino Acid ◽  
Brush Border ◽  
Amino Acid Transport ◽  
Brush Border Membrane ◽  
Membrane Vesicles ◽  
Brush Border Membrane Vesicles ◽  
Acid Transport ◽  
Transport Activity ◽  
Renal Brush Border Membrane ◽  
Renal Brush Border

Amino acid transport activity from bovine renal brush-border membrane vesicles (BBMV) was reconstituted into phospholipid vesicles composed of phosphatidylcholine/5% stearylamine. Reconstitutable transport activity was enhanced in protein fractions binding to various lectins. When solubilized BBMV were fractionated on peanut lectin, a single protein band of average molecular mass 132 kDa was obtained. When this protein fraction was reconstituted into phospholipid membrane vesicles, amino acid transport activity was obtained with properties similar to those in native BBMV with regard to amino acid specificity, although the cation specificity was different. A monoclonal antibody which reacted with the same protein removed reconstitutable amino acid transport activity from solubilized BBMV. These findings may provide the first identification of a renal amino acid-transporting protein, although confirmation of this identification by other approaches will be required.

scholarly journals A rapid method for the reconstitution of Na+-dependent neutral amino acid transport from bovine renal brush-border membranes

1987 ◽  
Vol 244 (3) ◽  
pp. 503-508 ◽  
Author(s):  
A M Lynch ◽  
J D McGivan
Keyword(s):  
Amino Acid ◽  
Rapid Method ◽  
Brush Border ◽  
Amino Acid Transport ◽  
Neutral Amino Acid ◽  
Acid Transport ◽  
Transport Activity ◽  
Brush Border Membranes ◽  
Renal Brush Border Membranes ◽  
Renal Brush Border

1. A simple and rapid method for the reconstitution of Na+-dependent neutral amino acid transport activity from bovine renal brush border membranes is described. 2. The neutral detergent decanoyl-N-methylglucamide (‘MEGA-10’) was employed to solubilize the membrane protein. This obviated the necessity for a prolonged dialysis step. 3. The properties of amino acid transport in these vesicles were similar to those observed in native membranes. 4. This should be a useful procedure in the eventual identification and isolation of amino acid transport proteins.

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