Is intestinal peptide transport energized by a proton gradient?

1985 ◽  
Vol 249 (2) ◽  
pp. G153-G160 ◽  
Author(s):  
Vadivel Ganapathy ◽  
Frederick H. Leibach

Transport of intact peptides, followed by intracellular hydrolysis in the intestinal mucosal cells, plays an important role in the absorption of protein digestion products in the mammalian small intestine. Even though earlier studies on peptide absorption in intact-tissue preparations have indicated that peptides are transported by an active Na+-dependent mechanism, recent studies with purified brush-border membrane vesicles have unequivocally demonstrated that Na+ does not play a direct role in the translocation of peptides across the membrane. Like most amino acids, peptides are also transported as zwitterions. However, peptide transport causes depolarization of the brushborder membrane in intact mucosal cells as well as in purified membrane vesicles, and the depolarization is the result of a net transfer of positive charge across the membrane during peptide transport. This electrogenic nature of peptide transport is observed even in the absence of Na+. Peptide transport is enhanced by an interior-negative membrane potential and inhibited by an interior-positive membrane potential. An inward proton gradient stimulates peptide transport, and this stimulation is reduced when the proton gradient is subjected to rapid dissipation by the presence of a proton ionophore. These observations strongly suggest that peptides are cotransported with protons in the intestine. There is substantial evidence for the existence of an inward proton gradient in the mammalian small intestine, and therefore it is very likely that this proton gradient is the in vivo energy source for the uphill transport of peptides. The Na+-H+ exchanger in the brush-border membrane, in conjunction with Na+-K+-ATPase at the basolateral membrane, is probably responsible for the generation and maintenance of the proton gradient and may thus be involved indirectly in the intestinal absorption of peptides.

1990 ◽  
Vol 269 (3) ◽  
pp. 565-571 ◽  
Author(s):  
J Dyer ◽  
R B Beechey ◽  
J P Gorvel ◽  
R T Smith ◽  
R Wootton ◽  
...  

The properties of a peptide-transport system in rabbit enterocyte basolateral membrane were examined with glycyl-L-proline as the substrate. Basolateral-membrane vesicles prepared from rabbit proximal intestine were characterized in terms of both purity and orientation. Marker-enzyme assays show that the basolateral-membrane marker, ouabain-sensitive K(+)-activated phosphatase, is enriched 17-fold with respect to the initial homogenate. The activities of enzymes used as markers for other membranes and organelles are low, and contamination of the final membrane fraction with these is minimal. The use of immunoblotting techniques further confirms the absence of brush-border-membrane contamination. Proteins in the basolateral-membrane vesicle preparation gave no cross-reaction with antibodies against the 140 kDa antigen and the Na+/glucose-symport protein, markers specific to the brush-border membrane of the enterocyte. Conversely, antibodies raised against the classical basolateral-membrane marker, the RLA class I histocompatibility complex, reacted strongly with a 43 kDa basolateral-membrane protein. The orientation of the basolateral-membrane vesicles was shown to be predominantly inside-out on determination by two independent criteria. The uptake of [1-14C]glycyl-L-proline by these vesicles is stimulated by the presence of an inwardly directed pH gradient, and this stimulation can be abolished by the proton ionophores carbonyl cyanide p-trichloromethoxyphenylhydrazone (CCCP) and tetrachlorotrifluoromethylbenzimidazole (TTFB). Transport is also inhibited by HgCl2, thimerosal, Na+ and other glycyl dipeptides.


1996 ◽  
Vol 199 (3) ◽  
pp. 635-641 ◽  
Author(s):  
M Thamotharan ◽  
G Ahearn

Epithelial brush-border membrane vesicles (BBMVs) of lobster (Homarus americanus) hepatopancreas were formed by a Mg2+ precipitation technique. In these BBMVs, [14C]glycylsarcosine ([14C]Gly-Sar) uptake was stimulated by a transmembrane proton gradient. transmembrane K+ diffusion potential (inside negative) stimulated [14C]Gly-Sar uptake above that observed with short-circuited vesicles, while an inwardly directed Na+ gradient had no stimulatory effect on peptide uptake. [14C]Gly-Sar influx (over 10 s) occurred by a low-affinity, saturable, proton-gradient-dependent carrier system (Kt=5.90±0.13 mmol l-1, Jmax=4662±487 pmol mg-1 protein 10 s-1; mean ± s.e.m., N=3). This carrier exhibited a high-affinity proton binding site (KH=235±25 nmol l-1; pK=6.6) and an apparent 1H+:1Gly-Sar transport stoichiometry. Influx of 0.1 mmol l-1 [14C]Gly-Sar into lobster hepatopancreatic BBMVs was significantly (P<0.01) cis-inhibited by 10 mmol l-1 diethylpyrocarbonate and by a variety of other dipeptides (10 mmol l-1), suggesting a broad transport specificity. These observations strongly suggest that transport of peptides into crustacean hepatopancreas is proton-gradient-dependent and electrogenic, qualitatively resembling the peptide transport paradigm proposed for fish and mammals.


1985 ◽  
Vol 249 (5) ◽  
pp. G607-G613 ◽  
Author(s):  
C. M. Schron ◽  
R. G. Knickelbein ◽  
P. S. Aronson ◽  
J. Della Puca ◽  
J. W. Dobbins

In the presence of a pH gradient (7.7 inside, 5.5 outside), the initial velocity of SO4 uptake by rabbit ileal brush-border membrane (BBM) vesicles was markedly stimulated compared with uptake in the absence of a pH gradient. Under pH gradient conditions, SO4 was transiently accumulated at a concentration 13-fold higher than at equilibrium ("overshoot"). Superimposition of a HCO3 gradient did not further stimulate the initial velocity of SO4 uptake compared with a pH gradient alone. Evidence that this pH gradient-stimulated SO4 uptake represented SO4-OH exchange included lack of sensitivity of SO4 transport to alterations of the membrane potential; 85-95% inhibition of SO4 uptake by the anion exchange inhibitors DIDS and SITS; and saturation kinetics (Km for SO4 = 0.475 +/- 0.054 mM; Vmax = 4.1 +/- 0.1 nmol SO4 X mg prot-1 X min-1). Sulfate did not inhibit pH gradient-stimulated 36Cl uptake, indicating that SO4-OH and Cl-HCO3(OH) are different exchangers. When BBM vesicles were compared with basolateral membrane (BLM) vesicles, pH gradient-stimulated SO4 uptake was found predominantly in the BBM preparation. Brush-border SO4-OH exchange was further localized by demonstrating Na-stimulated SO4 efflux from vesicles loaded under pH gradient conditions, suggesting that Na-SO4 cotransport and SO4-OH exchange are on the same BBM vesicles. In conclusion, a SO4-OH exchanger (or H-SO4 cotransporter) exists on the brush border of rabbit ileum which is distinct from the brush-border Cl-HCO3(OH) exchanger.(ABSTRACT TRUNCATED AT 250 WORDS)


1987 ◽  
Vol 252 (2) ◽  
pp. G281-G286 ◽  
Author(s):  
V. M. Rajendran ◽  
J. M. Harig ◽  
K. Ramaswamy

A proton-peptide symport mechanism has been postulated for transport of dipeptides in rabbit intestinal and renal brush-border membrane vesicles (BBMV). We have investigated the effects of a transmembrane potential (in mouse) and an inwardly directed proton gradient (in mouse, rabbit, and human) on the transport of glycyl-L-proline in intestinal BBMV. Membrane potential alterations, induced by permeant anions or generated by a K+-diffusion potential in the presence of valinomycin, did not accelerate the uptake of glycyl-L-proline. In contrast, in parallel experiments the uptake of D-glucose, whose cotransport system is electrogenic, was markedly increased by an interior negative membrane potential. Thus the transport of glycyl-L-proline in mouse intestinal BBMV is not electrogenic. Further studies on the effect of a proton gradient (extravesicular pH 5.5; intravesicular pH 7.5) on transport of glycyl-L-proline revealed an absence of stimulation of glycyl-L-proline transport and lower uptake rates in the presence of a proton gradient. The simultaneous presence of an interior negative membrane potential and an inwardly directed proton gradient did not accelerate the transport of glycyl-L-proline. These results demonstrate that the transport of glycyl-L-proline in mouse intestinal BBMV is neither electrogenic nor energized by an inwardly directed proton gradient. Likewise, pH gradients do not stimulate glycyl-L-proline uptake in either rabbit or human BBMV.


2021 ◽  
Vol 139 ◽  
pp. 109940
Author(s):  
Lesbia Cristina Julio-Gonzalez ◽  
F. Javier Moreno ◽  
María Luisa Jimeno ◽  
Elisa G. Doyagüez ◽  
Agustín Olano ◽  
...  

1981 ◽  
Vol 240 (3) ◽  
pp. F185-F191 ◽  
Author(s):  
E. F. Boumendil-Podevin ◽  
R. A. Podevin

The transport of nicotinic acid was investigated in brush border membrane vesicles isolated from rabbit kidney. The imposition of a Na+ gradient (out to in) induced a transient stimulation of nicotinic acid uptake above its final equilibrium value. This stimulation was specific for Na+. The uptake of nicotinic acid by the brush border membranes represented transport into an internal space and occurred in the absence of significant nicotinic acid degradation. The Na+ gradient-dependent uptake of nicotinic acid was saturable, apparent Km = 0.3 mM. Uptake of nicotinic acid was inhibited by its two isomers: picolinic and isonicotinic acid. In contrast, pyridine derivatives with two carboxyl groups or an amide group in addition to the carboxyl group were without inhibitory effect. Evaluation of changes in membrane potential using the lipophilic cation triphenylmethylphosphonium demonstrated that conditions that transiently generated either an interior-positive or an interior-negative membrane potential failed to affect the Na+-dependent transport of nicotinic acid. These findings provide evidence of the existence on the luminal membrane of a Na+ gradient-dependent and electroneutral transport system for nicotinic acid.


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