Interaction of the Insecticidal Crystal Protein CryIA from Bacillus thuringiensis with Amino Acid Transport into Brush Border Membranes from Bombyx mori Larval Midgut

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.

Download Full-text

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

Biochemical Journal ◽

10.1042/bj2900059 ◽

1993 ◽

Vol 290 (1) ◽

pp. 59-65 ◽

Cited By ~ 24

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.

Download Full-text

Amino acid transport systems in intestinal brush-border membranes from lepidopteran larvae

AJP Regulatory Integrative and Comparative Physiology ◽

10.1152/ajpregu.1989.257.3.r494 ◽

1989 ◽

Vol 257 (3) ◽

pp. R494-R500 ◽

Cited By ~ 1

Author(s):

B. Giordana ◽

V. F. Sacchi ◽

P. Parenti ◽

G. M. Hanozet

Keyword(s):

Amino Acids ◽

Amino Acid ◽

Brush Border ◽

Amino Acid Transport ◽

Transport Systems ◽

Acid Transport ◽

Specific System ◽

Neutral Amino Acids ◽

Intestinal Brush Border ◽

Lepidopteran Larvae

Experiments with intestinal brush-border membrane vesicles from lepidopteran larvae disclosed the occurrence of unique cotransporter proteins that use K+ as the driver cation for the transmembrane transfer of amino acids across the luminal border of midgut enterocytes. Six apical membrane amino acid transport systems have been identified. These systems are 1) a neutral amino acid transporter with a broad spectrum of interactions with most neutral amino acids, which is highly concentrative, strongly K+- and electrical potential-dependent, poorly stereospecific, and recognizes histidine, but not proline, glycine, or alpha-(methylamino)isobutyric acid (MeAIB); 2) a specific system for L-proline; 3) a specific system for glycine with a higher affinity for Na+ than for K+; 4) a specific system for L-lysine, which is dependent on membrane potential, is highly sensitive to external K+, and does not interact with L-arginine or neutral amino acids; 5) a specific K+-dependent process for glutamic acid, which does not recognize aspartic acid; and last, 6) an apparently unique K+- driven mechanism for D-alanine, which is potential-dependent and strongly stereospecific.

Download Full-text