Expression of Na+-d-glucose cotransporter in brush-border membrane of the chicken intestine

1999 ◽  
Vol 276 (2) ◽  
pp. R627-R631 ◽  
Author(s):  
Carles Garriga ◽  
Nativitat Rovira ◽  
Miquel Moretó ◽  
Joana M. Planas
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Western Blot ◽  
Brush Border ◽  
Synthetic Peptide ◽  
Brush Border Membrane ◽  
Polyclonal Antibodies ◽  
Membrane Vesicles ◽  
Dependent Transport

We have studied the expression of Na+-d-glucose cotransporter in brush-border membrane vesicles (BBMVs) of chicken enterocytes to correlate the changes in the apical Na+-dependent transport with the changes in the amounts of transporter determined by Western blot analysis. Two different rabbit polyclonal antibodies were used simultaneously. The antibody raised against amino acids 564–575 of the deduced amino acid sequence of rabbit intestinal SGLT-1 ( antibody 1) specifically detects a single 75-kDa band in the three segments, and this band disappeared when the antibody was preabsorbed with the antigenic peptide. The antibody raised against the synthetic peptide corresponding to amino acids 402–420 of the same protein ( antibody 2) only reacts with jejunal and ileal samples, but no signal is found in BBMVs of rectum. Only when antibody 1 was used was there a linear correlation between the maximal transport rates of hexoses in BBMVs and the relative protein amounts determined by Western blot. These results indicate that the Na+-d-glucose cotransport in the jejunum, the ileum, and the rectum of chickens is due to an SGLT-1 type protein.

Potential differences influence amino acid/Na+ symport rates in larval Manduca sexta midgut brush-border membrane vesicles.

1994 ◽  
Vol 189 (1) ◽  
pp. 55-67
Author(s):  
R Parthasarathy ◽  
W R Harvey
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Manduca Sexta ◽  
Brush Border ◽  
Brush Border Membrane ◽  
Membrane Vesicles ◽  
Brush Border Membrane Vesicles ◽  
Rate Determining Step ◽  
Half Saturation Constant

The time-dependent fluorescence intensity of an intravesicular potential-sensitive dye was used to probe the real-time kinetics of potential difference (PD)-dependent amino acid/Na+ symport at pH9 into brush-border membrane vesicles obtained from larval Manduca sexta midgut. Neutral amino acids (alanine, proline) are symported at higher rates as the vesicles are hyperpolarized. The symport rates of acidic (glutamate) and basic (arginine) amino acids are almost PD-independent. The half-saturation constant of alanine is PD-independent between -108 and -78 mV, although the maximal symport velocity increases by half as the voltage is increased. Amino acid throughput is evidently enhanced as the relatively high transmembrane PDs (> 150 mV, lumen positive) measured in vivo are approached. The half-saturation concentrations of Na+ were in the range 15-40 mmol l-1 for most of the amino acids examined and increased with voltage for alanine. The Vmax observed as a function of cation or amino acid concentration increased as the vesicle was hyperpolarized in the case of leucine and alanine. The data support the hypothesis that carrier and substrates are at equilibrium inasmuch as substrate translocation seems to be the rate-determining step of symport.

scholarly journals Characterization of tryptophan transport in human placental brush-border membrane vesicles

1986 ◽  
Vol 238 (1) ◽  
pp. 201-208 ◽  
Author(s):  
M E Ganapathy ◽  
F H Leibach ◽  
V B Mahesh ◽  
J C Howard ◽  
L D Devoe ◽  
...  
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Brush Border ◽  
Brush Border Membrane ◽  
Aromatic Amino Acids ◽  
Membrane Vesicles ◽  
Brush Border Membrane Vesicles ◽  
System 1 ◽  
Tryptophan Uptake ◽  
Tryptophan Transport

The characteristics of tryptophan uptake in isolated human placental brush-border membrane vesicles were investigated. Tryptophan uptake in these vesicles was predominantly Na+-independent. Uptake of tryptophan as measured with short incubations occurred exclusively by a carrier-mediated process, but significant binding of this amino acid to the membrane vesicles was observed with longer incubations. The carrier-mediated system obeyed Michaelis-Menten kinetics, with an apparent affinity constant of 12.7 +/- 1.0 microM and a maximal velocity of 91 +/- 5 pmol/15 s per mg of protein. The kinetic constants were similar in the presence and absence of a Na+ gradient. Competition experiments showed that tryptophan uptake was effectively inhibited by many neutral amino acids except proline, hydroxyproline and 2-(methylamino)isobutyric acid. The inhibitory amino acids included aromatic amino acids as well as other system-1-specific amino acids (system 1 refers to the classical L system, according to the most recent nomenclature of amino acid transport systems). The transport system showed very low affinity for D-isomers, was not affected by phloretin or glucose but was inhibited by p-azidophenylalanine and N-ethylmaleimide. The uptake rates were only minimally affected by change in pH over the range 4.5-8.0. Tryptophan uptake markedly responded to trans-stimulation, and the amino acids capable of causing trans-stimulation included all amino acids with system-1-specificity. The patterns of inhibition of uptake of tryptophan and leucine by various amino acids were very similar. We conclude that system t, which is specific for aromatic amino acids, is absent from human placenta and that tryptophan transport in this tissue occurs via system 1, which has very broad specificity.

A novel proline, glycine: K+ symporter in midgut brush-border membrane vesicles from larval Manduca sexta.

1995 ◽  
Vol 198 (12) ◽  
pp. 2599-2607 ◽  
Author(s):  
A L Bader ◽  
R Parthasarathy ◽  
W R Harvey
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Manduca Sexta ◽  
Brush Border ◽  
Brush Border Membrane ◽  
Membrane Vesicles ◽  
Brush Border Membrane Vesicles ◽  
Neutral System ◽  
Aminoisobutyric Acid ◽  
Posterior Midgut

Alkali-cation-dependent uptake of proline and glycine into brush-border membrane vesicles from the midgut of the larval tobacco hornworm Manduca sexta was investigated using rapid filtration assays. Uptake of both amino acids was by electrophoretic symport, with K+ being the favored cation at pH 10. Counterflow accumulation of proline was elicited by glycine and vice versa, suggesting that the two amino acids are transported by a common symporter, which we designate the pro, gly: K+ symporter. L-alpha-Aminoisobutyric acid was the only other amino acid that elicited the accumulation of both proline and glycine. D-Proline was not symported; L-proline, glycine and L-alpha-aminoisobutyric acid appear to be the only substrates of the pro, gly: K+ symporter. Neutral amino acids with relatively short sidechains elicit glycine accumulation, suggesting that glycine may also be symported by the well-established neutral amino acid system. Since proline does not utilize the broad-spectrum, neutral system, its symport appears to be exclusively through the pro, gly: K+ symporter. Proline symport was found mainly in posterior midgut vesicles, suggesting that the pro, gly: K+ symporter may be localized in this region of the midgut.

AMINO ACID TRANSPORT SYSTEMS IN BRUSH-BORDER MEMBRANE VESICLES FROM LEPIDOPTERAN ENTEROCYTES

1989 ◽  
Vol 143 (1) ◽  
pp. 87-100
Author(s):  
GIORGIO M. HANOZET ◽  
BARBARA GIORDANA ◽  
V. FRANCA SACCHI ◽  
PAOLO PARENTI
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Glutamic Acid ◽  
Brush Border ◽  
Amino Acid Transport ◽  
Brush Border Membrane ◽  
Membrane Vesicles ◽  
Transport Systems ◽  
Acid Transport ◽  
Luminal Membrane

The presence of different potassium-dependent amino acid transport systems in the luminal membrane of the larval midgut of Philosamia cynthia Drury (Saturnidae, Lepidoptera) was investigated by means of countertransport experiments performed with brush-border membrane vesicles. The vesicles were preloaded with 14 different unlabelled amino acids, whose ability to elicit an intravesicular accumulation over the equilibrium value of six labelled amino acids (L-alanine, L-leucine, L-phenylalanine, L-glutamic acid, L-lysine and L-histidine) was tested. For histidine, the results were compared with those obtained from inhibition experiments, in which the same 14 amino acids were used as inhibitors on the cis side of the brush-border membrane. The data demonstrate the presence in the lepidopteran luminal membrane of distinct transport pathways for lysine and glutamic acid. The transport of most neutral amino acids, with the exclusionof glycine and proline, seems to occur through a system that may be similar to the neutral brush-border system (NBB) found in mammalian intestinal membranes. This system is also able to handle histidine.

scholarly journals Electroneutral Na+-2 Cl−-Leucine Cotransport by Lobster Hepatopancreatic Brush-Border Membrane Vesicles

1988 ◽  
Vol 136 (1) ◽  
pp. 363-381
Author(s):  
GREGORY A. AHEARN ◽  
LAUREL P. CLAY
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Brush Border ◽  
Brush Border Membrane ◽  
Membrane Vesicles ◽  
Brush Border Membrane Vesicles ◽  
Acidic Ph ◽  
Polar Species ◽  
L System ◽  
Leucine Transport

Uptake of L-[3H]leucine by lobster hepatopancreatic brush-border membrane vesicles was stimulated by a transmembrane NaCl gradient (o>i), but not by identical gradients of NaSCN or other Cl− salts (e.g. K+, Li+, NH4+, Cs+ or choline), suggesting that amino acid transfer depended upon both Na+ and Cl−. In NaCl medium at acidic pH, leucine uptake was largely electroneutral and unresponsive to a transmembrane potential generated by permeable anions; however, in Na+-free medium, amino acid transport was strongly electrogenic under the same conditions. Leucine influx occurred by a combination of two carrier processes at physiologically acidic pH. One exhibited an influx Kt of 0.59 mmol l−1, a JM of 390 pmol mg protein−1 S−1 and a cotransport stoichiometry of 1 Na+: 2 Cl+: 1 leucine. This process was most strongly cis-inhibited by the non-polar amino acids phenylalanine, methionine and isoleucine, and most weakly inhibited by the more polar species methylaminoisobutyric acid (MeAIB), hydroxyproline, glutamate and arginine. The second leucine carrier system showed a very low binding affinity and could not be distinguished from diffusion, was Na+- and Cl−-independent, and was cis-inhibited by more polar amino acids such as lysine, hydroxyproline, MeAIB, alanine and glutamate. These results suggest that brush-border leucine transport in lobster hepatopancreas at acidic pH may occur by a combination of a modified L-system, that includes ion cosubstrates, and either by a second undefined Na+-independent process with a broad structural specificity or by multiple Na+-independent processes.

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