Pteroylpolyglutamate hydrolase from human jejunal brush borders. Purification and characterization

Pteroylpolyglutamate hydrolase was solubilized with Triton X-100 from human jejunal mucosal brush borders and purified approximately 5,000-fold using organomercurial affinity chromatography, DEAE-cellulose chromatography, and gel filtration. The apparent molecular weight of the purified enzyme in the Triton micelle was estimated as 700,000 using Bio-Gel A-1.5m gel filtration. Sodium dodecyl sulfate/urea-polyacrylamide gel electrophoresis followed by Coomassie stain demonstrated two polypeptide bands at 145,000 and 115,000 daltons. The purified enzyme had an isoelectric point of 7.2, was maximally active at pH 5.5, and was stable above pH 6.5 and at temperatures up to 65 degrees C for at least 90 min. Human jejunal brush-border pteroylpolyglutamate hydrolase is an exopeptidase which liberated [14C]Glu as the sole labeled product of PteGlu2[14C]Glue (where PteGlun represents pteroylpolyglutamate), failed to liberate a radioactive product from PteGlu2[14C]GluLeu2, and released all possible labeled PteGlun products during incubation with Pte[14C]GluGlu6 with the accumulation of Pte[14C]Glu. PteGlu2, PteGlu3, and PteGlu7 were substrates, each with Km = 0.6 microM, whereas PteGlu was a weak inhibitor of the hydrolysis of PteGlu3 with Ki = 20 microM. Components of the pteroyl moiety, Glu, and short chain Glun in alpha or gamma linkages were not inhibitory. The enzyme was activated by Zn2+ or Co2+. The properties of brush-border pteroylpolyglutamate hydrolase are different from those described for the soluble intracellular pteroylpolyglutamate hydrolase in other species and in human mucosa, yet are consistent with previous data on the process of hydrolysis of PteGlun in the intact human intestine.     

Jejunal brush-border folate hydrolase. A novel enzyme.

Dietary folate, a vitamin required for DNA synthesis and cell regeneration, occurs as pteroylpolyglutamates that are hydrolyzed to pteroylglutamate during the process of intestinal absorption. Studies from our laboratory over the past 15 years have shown that jejunal brush-border folate hydrolase is essential and rate-limiting in folate absorption. Brush-border folate hydrolase activity and pteroylpolyglutamate hydrolysis are inhibited in disease and conditions associated with folate deficiency, including celiac and tropical sprue, the use of sulfasalazine to treat inflammatory bowel disease, and chronic alcoholism. Brush-border folate hydrolase is an exopeptidase located on the jejunal brush-border surface that liberates hydrolytic products of pteroylpolyglutamates in a progressive fashion, with a final release of pteroylglutamate. Subsequent steps in folate absorption include uptake by a brush-border folate-binding-protein receptor and transport across the brush-border membrane into the enterocyte. These steps are probably followed by an intracellular synthesis of pteroylglutamates for folate-dependent reactions and intracellular hydrolysis to pteroylglutamate for transport across the basolateral membrane to the portal circulation. In pigs, the active form of jejunal brush-border folate hydrolase has a molecular weight of 240 kd and is probably a homodimer of the 120-kd protein found after immunoprecipitation with specific antibody. Regulating the synthesis and expression of brush-border folate hydrolase may be critical to the availability of dietary folate.     

Some comparative properties and localization of porcine jejunal adenylate cyclase.

Cholera toxin is thought to cause intestinal secretion by activating adenylate cyclase and increasing intracellular 3',5'-cyclic AMP concentrations in intestinal mucosa. Cholera toxin causes profuse secretion of fluid into ligated intestinal loops of both pigs and rabbits, but cholera toxin-induced increases in 3',5'-cyclic AMP concentration are much lower in the pig than in the rabbit. Porcine jejunal adenylate cyclase was examined for unusual properties which might account for a lack of 3'-5'-cyclic AMP accumulation after treatment with cholera toxin. The divalent cation requirements, the pH optimum, and the stimulation by fluoride ion were unremarkable. The Km for ATP was 0.11 mM with negative cooperativity indicated by a Hill coefficient of 0.83. Triton X-100 was inhibitory and guanosine diphosphate methylenephosphate stimulated enzyme activity. Adenylate cyclase activity was highest in the basal and lateral membrane fractions of jejunal mucosa and relatively low in brush-border preparations. Pretreatment of pig jejunum with cholera toxin caused a 30-40% activation of the crude and of the partly purified enzyme. A relatively low activation of adenylase cyclase in pig jejunal mucosa, compared with rabbit, may account for the absence of 3',5'-cyclic AMP accumulation after cholera-toxin treatment in the pig.     

Similarities between a dipeptide hydrolase from brush-border and cytosol fractions of guinea-pig intestinal mucosa.

A dipeptide hydrolase from the brush border of guinea-pig intestinal mucosa was purified. The enzyme resembles another dipeptide hydrolase isolated from the cytosol fraction of intestinal mucosa. Studies on the binding of cytosol peptide hydrolase to brush-border membranes indicate that the enzyme found in the brush border may be a cytoplasmic contaminant.     

Human jejunal brush border folate conjugase. Characteristics and inhibition by salicylazosulfapyridine

Human jejunal brush border folate conjugase (EC 3.4.22.-) was partially purified and characterized. Three drugs known to be associated with clinical folate deficiency were tested for inhibition of the partially purified enzyme. Using jejunal mucosa from obese patients undergoing intestinal bypass surgery, brush border folate conjugase was purified 50-80-fold by centrifugation, Triton X-100 solubilization and DEAE-Sephadex and Sephacryl S-200 chromatography. Using synthetic pteroyldiglutamyl[14C]glutamate as substrate, the enzyme was found to have a pH optimum of 6.5 and an apparent Km of 1.6 micro M. Incubation of the enzyme with synthetic pteroyl[14C]glutamylhexaglutamate resulted in a spectrum of shorter-chain 14C-labeled pteroylglutamates at 60 min. Pteroyl[14C]glutamate was the major product at 120 min, with quantitative recovery of free glutamate in the incubation medium. Salicylazosulfapyridine was a competitive inhibitor of the enzyme (Ki = 0.13 mM), while ethanol, diphenylhydantoin and salicylazosulfapyridine metabolites had no effect. These data suggest that brush border folate conjugase is an exopeptidase which progressively hydrolyzes glutamyl units from pteroylpolyglutamate, leaving pteroylmonoglutamate as the folate form available for intestinal transport. Inhibition of brush border folate conjugase by salicylazosulfapyridine provides a mechanism for folate malabsorption and deficiency in chronic users of this drug.     

Transport characteristics of L-glutamate in human jejunal brush-border membrane vesicles

Previous work using human jejunal brush-border membrane vesicles has demonstrated the existence of a distinct transport system in man for acidic amino acids. This system is energized by an inwardly directed Na+ gradient and an outwardly directed K+ gradient. These studies further characterize the transport of L-glutamate in the human jejunal brush-border membrane vesicles. Efflux studies were performed by loading the brush-border membrane vesicles with radiolabeled L-glutamate and sodium chloride. Extravesicular K+ accelerated the efflux of L-glutamate when compared to extravesicular Na+ or choline, indicating that potassium serves to recycle the carrier. Unlabeled extravesicular L-glutamate (but not D-glutamate) also enhanced the efflux of radiolabeled L-glutamate demonstrating that there is a bidirectional similarity to the transport system. The effect of pH on the transport system was also investigated by varying the intravesicular and extravesicular pH from 5.5 to 9. A pH environment of 6.5 produced the highest initial uptake rates as well as the greatest overshoots for transport of L-glutamate into brush-border membrane vesicles. The imposition of an inwardly directed pH gradient (5.5 outside, 7.5 inside) accelerated both the influx and efflux of L-glutamate. These results demonstrate that the L-glutamate carrier system in human jejunum appears to have similar energizing characteristics in either direction across the brush-border membrane. In addition, the system operates at an optimal pH of 6.5 and protonation of the system may enhance its mobility.     

Cholesterol ester hydrolase(s) in mammalian brain: Is there a myelin-specific cholesterol ester hydrolase?

The present study compared the properties of cholesterol ester hydrolase(s) in myelin and microsomes from rat, mouse and human brain. The results indicated that the enzyme activity in both myelin and microsomes from rat, mouse and human brain was optimal at pH 6.5 and required Triton X-100 for optimal activity. The enzyme activity in myelin was 3- to 4-fold higher in the presence of Trition X-100 than taurocholate. Addition of phosphatidyl serine enhanced (2 to 4 fold) the hydrolase activity in both myelin and microsomes. The properties of the enzyme in solubilized preparation of myelin were also similar to the properties of the enzyme in partially delipidated and solubilized preparations of microsomes. The activity was again optimal at pH 6.5, required Triton X-100 for optimal activity and was stimulated by phosphatidyl serine. These results indicate that the properties of cholesterol ester hydrolase in myelin are similar to those of the microsomal enzyme and that this is true for the fractions from both human and rodent brain. The data thus lead us to believe that the hydrolase activity in mammalian brain myelin and microsomes may reflect the distribution of a single enzyme in the two fractions rather than two distinct enzymes, one being specific to each fraction.     

Isolation and characterization of arginine ester hydrolase from Heloderma horridum (beaded lizard) venom.

1. An arginine ester hydrolase was isolated from Heloderma horridum (beaded lizard) venom by Sephadex G-75, DEAE-Sephacel and Q-Sepharose column chromatography, resulting in 5.4 mg of purified enzyme from 320.0 mg of crude venom. 2. The enzyme was shown to be homogeneous by both SDS and non-SDS disc electrophoresis on polyacrylamide gel at pH 8.3. 3. The enzyme possesses arginine ester hydrolase and transglutaminase-like activities, but did not exhibit clotting activity. 4. Molecular weight was determined to be ca 29 kDa, with an isoelectric point of 4.4. 5. The enzyme was stable to heat treatment (95 degrees C, 10 min) and to pH changes over the range 2-11. 6. The arginine ester hydrolase was inactivated by diisopropylfluorophosphate (DFP), beta-mercaptoethanol and N-bromosuccinimide, suggesting that serine, disulfide bonds and tryptophan are involved in enzymatic activity. 7. Amino terminal sequences were determined and appear to be similar to porcine pancreatic kallikrein.     

Location of the two catalytic sites in intestinal lactase-phlorizin hydrolase. Comparison with sucrase-isomaltase and with other glycosidases, the membrane anchor of lactase-phlorizin hydrolase.

Lactase-phlorizin hydrolase was isolated by immunoadsorption chromatography from rabbit brush-border membrane vesicles. Inactivation of the enzyme with [3H]conduritol-B-epoxide, a covalent active site-directed inhibitor, labeled glutamates at positions 1271 and 1747. Glu1271 was assigned to lactase, Glu1747 to phlorizin hydrolase activity. In contrast, the nucleophiles in the active sites of sucrase-isomaltase are aspartates (Asp505 and Asp1394). Asp505 is a part of the isomaltase active site and is localized on the larger subunit, which carries the membrane anchor also, while Asp1394 is a part of the active of sucrase. Alignment of these 2 nucleophilic Glu residues in lactase-phlorizin hydrolase and of their flanking regions with published sequences of several other beta-glycosidases allows the classification of the configuration retaining glycosidases into two major families: the "Asp" and the "Glu" glycosidases, depending on the carboxylate presumed to interact with the putative oxocarbonium ion in the transition state. We offer some predictions as to the Glu acting as the nucleophile in the active site of some glycosidases. By hydrophobic photolabeling, the membrane-spanning domain of lactase-phlorizin hydrolase was directly localized in the carboxyl-terminal region thus confirming this enzyme as a monotopic type I protein (i.e. with Nout-Cin orientation) of the brush-border membranes. A simplified version of the Me2+ precipitation method to efficiently and simply prepare brush-border membrane vesicles is also reported.     

Purification and characterization of alpha-L-fucosidase from Streptomyces species.

Streptomyces sp. 142, isolated from a soil sample, produced alpha-fucosidase when cultured in the presence of L-fucose. The enzyme was purified 700-fold with an overall recovery of 17% from a cell-free extract by cation exchange chromatography and gel filtration chromatography. The apparent molecular weight of the purified enzyme was 40,000 by gel filtration chromatography. The enzyme had a pH optimum of 6.0 and was stable at pH 4.5-7.0. Substrate specificity studies with oligosaccharides labeled with 2-aminopyridine as the substrate showed that the enzyme specifically hydrolyzed terminal alpha 1-3 and alpha 1-4 fucosidic linkages in the oligosaccharides but did not hydrolyze alpha 1-2 or alpha 1-6 fucosidic linkages or a synthetic substrate, p-nitro-phenyl alpha-L-fucoside. The purified enzyme released L-fucose from a fucosylated glycoprotein, alpha 1-acid glycoprotein. Thus, the substrate specificities of the Streptomyces alpha-fucosidase resembled those of alpha-fucosidases I and III isolated from almond emulsin rather than those of other microbial alpha-fucosidases.     

Characteristics of tripeptide transport in human jejunal brush-border membrane vesicles

These studies are aimed at characterizing the transport of the tripeptide, glycylglycyl-L-proline (GlyGlyPro) across human jejunal brush-border membrane vesicles. GlyGlyPro (0.65 mM) was hydrolyzed by brush-border membrane vesicles with the extent of hydrolysis per mg protein being 23% at 0.5 min, 57% at 1 min and complete hydrolysis at 60 min. Treatment of the membrane vesicles with gel-complexed papain (to remove membrane peptidases) resulted in minimal hydrolysis of GlyGlyPro up to 10 min of incubation. Measurement of GlyGlyPro influx with papain-treated vesicles in the presence of increasing medium osmolarity showed that uptake occurred into an osmotically reactive intravesicular space. Transport of GlyGlyPro with normal and papain-treated membrane vesicles was similar in the presence of an inward Na+ or K+ gradient. No overshoot phenomenon was observed in the presence of an inward proton gradient (extravesicular pH 5.5; intravesicular pH 7.5). An interior negative membrane potential induced by a K+ diffusion potential in the presence of valinomycin stimulated the uptake of the peptide. The effect of increasing concentrations on initial rates of GlyGlyPro uptake revealed the presence of a saturable component as well as a diffusional component. Preloading the membrane vesicles with 20 mM glycylsarcosylsarcosine stimulated uptake by 4-fold. Uptake of GlyGlyPro was inhibited greater than 50% by dipeptides and tripeptides and less than 15% by free amino acids. These results indicate that GlyGlyPro uptake in jejunal brush-border membrane vesicles is not energized by a Na+ or proton gradient and that transport occurs by carrier-mediated and diffusional processes.     

Aboral changes in D-glucose transport by human intestinal brush-border membrane vesicles.

D-Glucose transport was investigated in isolated brush-border membrane vesicles from human small intestine. Characteristics of D-glucose transport from the jejunum were compared with that in the mid and terminal ileum. Jejunal and mid-ileal D-glucose transport was Na+-dependent and electrogenic. The transient overshoot of jejunal D-glucose transport was significantly greater than corresponding values in mid-ileum. The terminal ileum did not exhibit Na+-dependent D-glucose transport, but did exhibit Na+-dependent taurocholate transport. Na+-glucose co-transport activity as measured by tracer-exchange experiments was greatest in the jejunum, and diminished aborally. We conclude that D-glucose transport in man is Na+-dependent and electrogenic in the proximal intestine and directly related to the activity of D-glucose-Na+ transporters present in the brush-border membranes. D-Glucose transport in the terminal ileum resembles colonic transport of D-glucose.     

Evidence for Two Cholesterol Ester Hydrolases in Human Cerebrospinal Fluid

Abstract: In the present study, the properties, such as pH optima, detergent requirement, and effects of various lipids, of cholesterol ester hydrolase in human cerebrospinal fluid (CSF) were examined, and the activity levels of the enzyme in CSF from multiple sclerosis (MS) patients and non-MS individuals were compared. Our data indicate that the enzyme in CSF exhibits two pH optima: pH 6.0 in the presence of Triton X-100 and pH 7.0 in the presence of sodium taurocholate. Both phosphatidylethanolamine (PE) and phosphatidylserine (PS) enhanced the hydrolase activity at pH 6.0. The activity at pH 7.0, on the other hand, was enhanced slightly in the presence of PE but was inhibited in the presence of PS. These data suggest the presence of two cholesterol ester hydrolases in CSF and also indicate that the activity at pH 6.0 may be due to microsomal enzyme in brain and that at pH 7.0 may be due to myelin enzyme. The hydrolase activity at pH 7.0 was significantly lower in CSF from MS patients. The activity at pH 6.0 in CSF from MS and non-MS patients, however, did not differ significantly. This indicates that the reduction in pH 7.0 hydrolase activity in CSF may be related to demyelination.     

Purification and Properties of a Glycerol Ester Hydrolase (Lipase) from Propionibacterium shermanii

An intracellular glycerol ester hydrolase (lipase) from Propionibacterium shermanii was recovered from cell-free extracts and purified by ammonium sulfate precipitation, gel filtration, and ion-exchange chromatography on diethylaminoethylcellulose. Maximum enzyme activity was observed at pH 7.2 and 47 C when an emulsion of tributyrin was used as substrate. The enzyme was stable between pH 5.5 and 8. Heating the enzyme solution at 45 C for 10 min resulted in a 75% decrease in activity. Maximum rate of hydrolysis of triglycerides was observed on tripropionin, followed in order by tributyrin, tricaproin, and tricaprylin. The lipase was strongly inhibited by mercury and arsenicals, but specific sulfhydryl reagents had little or no inhibiting effect on the enzyme activity. The enzyme also showed some esterase activity, but the hydrolysis of substrates in solution was small as compared to the hydrolysis of substrates in emulsion.     

Intracellular degradation of poly(3-hydroxybutyrate) granules of Zoogloea ramigera I-16-M.

Intracellular degradation of poly(3-hydroxybutyrate) (PHB) in bacteria is not yet clear. The properties of the autodigestion of native PHB granules from Zoogloea ramigera I-16-M were examined. The release of D(-)-3-hydroxybutyrate was observed only at pH values higher than about 8.5 and at relatively high ionic strength (optimal concentration 200 mM NaCl). Triton X-100 and diisopropylfluorophosphate inhibited this reaction. Addition of the supernatant fraction of Z. ramigera did not increase the release of D(-)-3-hydroxybutyrate from the native PHB granules. On the other hand, using the protease-treated PHB granules from Alcaligenes eutrophus as a substrate, PHB depolymerase activity was detected in the supernatant fraction of Z. ramigera cells. The soluble PHB depolymerase showed similar properties to the enzyme in the PHB granules. Since PHB depolymerase activity was found in fractions containing D(-)-3-hydroxybutyrate oligomer hydrolase activity, which were separated by DEAE-Toyopearl or by Sephacryl S-100, it is possible that the intracellular PHB depolymerase is identical to the oligomer hydrolase which has been purified already.     

Intracellular degradation of poly(3-hydroxybutyrate) granules of Zoogloea ramigera I-16-M

Abstract Intracellular degradation of poly(3-hydroxybutyrate) (PHB) in bacteria is not yet clear. The properties of the autodigestion of native PHB granules from Zooglea ramigera I-16-M were examined. The release of d (−)-3-hydroxybutyrate was observed only at pH values higher than about 8.5 and at relatively high ionic strength (optimal concentration 200 mM NaCl). Triton X-100 and diisopropylfluorophosphate inhibited this reaction. Addition of the supernatant fraction of Z. ramigera did not increase the release of d (−)-3-hydroxybutyrate from the native PHB granules. On the other hand, using the protease-treated PHB granules from Alcaligenes eutrophus as a substrate, PHB depolymerase activity was detected in the supernatant fraction of Z. ramigera cells. The soluble PHB depolymerase showed similar properties to the enzyme in the PHB granules. Since PHB depolymerase activity was found in fractions containing d (−)-3-hydroxybutyrate oligomer hydrolase activity, which were separated by DEAE-Toyopearl or by Sephacryl S-100, it is possible that the intracellular PHB depolymerase is identical to the oligomer hydrolase which has been purified already.     

Immunocytochemical localization of lysosomal beta-galactosidase in rat liver

beta-galactosidase is a ubiquitous lysosomal hydrolase that specifically cleaves terminal beta-galactosyl residues from glycoproteins, glycosaminoglycans, oligosaccharides, and glycolipids. To study the intracellular distribution of this enzyme, we prepared a specific polyclonal antibody to lysosomal beta-galactosidase by immunizing rabbits with a highly purified preparation of beta- galactosidase from rat liver. Using this antibody we employed an immunocytochemical technique (protein A coupled to horseradish peroxidase and diaminobenzidine cytochemistry) and showed that beta- galactosidase is present in all hepatocytes of the rat liver. All types of lysosomes, the rough endoplasmic reticulum, and the specialized region of smooth endoplasmic reticulum known as GERL showed immunoreactivity. This in situ distribution suggests that these organelles are involved in the biosynthesis and intracellular sorting of this lysosomal enzyme.     

Na+-dependent elevation of the acidic cell surface pH (microclimate pH) of rat jejunal villus cells induced by cyclic nucleotides and phorbol ester: possible mediators of the regulation of the Na+/H+ antiporter

The effects of cyclic nucleotides and phorbol ester on the acidic cell surface pH of rat jejunal villi were studied by using single-barrelled pH-sensitive microelectrodes. Addition of dibutyryl cAMP (1 mM) to the mucosal bathing solution caused an elevation of the cell surface pH from 6.19 +/- 0.04 (n = 12 measurements from three animals) to 6.53 +/- 0.03 (12) in the presence of Na+ in the medium. However, dibutyryl cAMP had no significant effect in the absence of Na+ and presence of 1 mM amiloride. Dibutyryl cGMP (1 mM) also had an Na+-dependent inhibitory effect on the cell surface pH. A phorbol ester, phorbol 12-myristate 13-acetate, caused an elevation of the cell surface pH only in the presence of Na+ from 6.14 +/- 0.07 (12) to 6.46 +/- 0.08 (12). Phorbol and phorbol 13-acetate, which do not stimulate protein kinase C, were without significant effects. These results suggest that increased levels of the intracellular cyclic nucleotides and activation of protein kinase C raise the acidic cell surface pH by inhibiting the activity of the brush-border Na+/H+ antiporter in the rat jejunal villus cells.     

Metabolism of resorcinylic compounds by bacteria. Purification and properties of acetylpyruvate hydrolase from Pseudomonas putida 01.

Acetylpyruvate hydrolase, the terminal inducible enzyme of the pathway of orcinol catabolism in Pseudomonas putida, catalyzes the quantitative conversion of acetylpyruvate into acetate and pyruvate. The enzyme has been purified approximately 40-fold from extracts of Ps. putida grown on orcinol. Disc gel electrophoresis of the preparations show one major and one minor band of protein. The molecular weight of the enzyme is approximately 38,000 by sodium dodecyl sulfate electrophoresis. Acetylpyruvate is the only known substrate for the enzyme; maleylpyruvate, fumarylpyruvate, acetoacetate, oxalacetate, and acetylacetone are not hydrolyzed by acetylpyruvate hydrolase. Several divalent cations, includ-Mg2+, Mn2+, Co2+, Ca2+, and Zn2+, enhanced hydrolytic activity, but Cu2+ was inhibitory. The enzyme shows a sharp pH optimum at 7.4. Acetylpyruvate hydrolase has an apparent K-m of 0.1 mM for acetylpyruvate with a molecular activity of 36 min minus 1 at 25 degrees. Pyruvate, oxalacetate, and oxalate are competitive inhibitors of acetylpyruvate hydrolysis by the enzyme with K-i values of 6.0, 4.5, and 0.45 mM, respectively.     

The effects of dietary phosphorus deficiency on surface pH and membrane composition of the mucosa epithelium in caprine jejunum

In ruminants, the uptake of inorganic phosphate (P_i) across the intestinal mucosa epithelium by Na-dependent and Na-independent mechanisms is a main regulatory factor in P homeostasis. The aim of the study was to elucidate to which extent Na-independent mechanisms, including pH effects or composition of mucosal brush-border membranes, could be involved in positive stimulation of P_i absorptive processes seen under the P deficient condition. Therefore, luminal, surface and intracellular pH of the jejunal epithelial cells in control and P depleted goats were compared and biochemical analyses of membrane phospholipids in the apical membrane of the jejunal epithelium were performed. Dietary P depletion resulted in decreased plasma P_i levels. While pH in jejunal ingesta was not significantly changed, P depletion resulted in a significantly lower surface pH in the crypt region compared to control animals (7.62 ± 0.02 vs. 7.77 ± 0.04, n = 4, P < 0.01). Inhibition of apical Na⁺/H⁺-exchange resulted in an increase of the jejunal surface pH in P depleted animals by 0.07 ± 0.01 (n = 6, P < 0.01) and 0.05 ± 0.01 (n = 6, P < 0.01) for the villus and the crypt region, respectively. This increase were inversely correlated with the initial surface pH prior to inhibition. In contrast to surface pH, intracellular pH of the jejunal epithelium and the phospholipid composition of the apical jejunal membrane were not affected by P depletion. Although the data suggest the existence of a Na⁺/H⁺-exchange mechanism at the luminal surface of goat jejunum they do not support the hypothesis that adaptational processes of active P_i absorption from goat jejunum in response to low dietary P could be based on “non P_i transporter events”.