Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.2.1.108 (lactase)
 2,133 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The biosynthesis of rat intestinal lactase-phlorizin hydrolase was studied by pulse-labeling of jejunal explants from 5-day-old suckling rats in organ culture. Explants were either continuously labeled with [35S] methionine for 15, 30, and 60 min or pulse-labeled for 30 min and chased for various periods of time up to 6 h in the presence or absence of protease inhibitors (PI), leupeptin, phenylmethylsulfonyl fluoride, and soybean trypsin inhibitor. Lactase-phlorizin hydrolase was immunoprecipitated from microvillus membrane (MVM) and ER-Golgi fractions with monoclonal antibodies. After pulse-labeling, lactase-phlorizin hydrolase from the ER-Golgi fraction appeared on SDS-PAGE as one band of approximately 220 kDa, regardless of the presence or absence of PI in the culture media. The 220-kDa protein band could also be labeled after incubation with [2-3H]mannose. In the absence of PI, the 220-kDa band appeared in the MVM by 30 min chase, simultaneously with a 180-kDa band, and by 60 min of chase an additional band of 130 kDa was seen. With increasing time of chase, the relative intensity of the 130-kDa band increased, whereas that of the 220-kDa band decreased, suggesting a precursor-product relationship. When PI were added to the medium, the formation of the 180-kDa band was not affected, but the conversion of the 180-kDa protein to the 130-kDa protein was virtually blocked. These findings suggest that lactase-phlorizin hydrolase is initially synthesized as a glycosylated precursor of 220 kDa, which is transported to the MVM. There it undergoes the following two cleavages: first, to the 180-kDa form, which is not prevented by PI used in these experiments, and second, to the 130-kDa form inhibited by PI.
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PMID:Biosynthesis, glycosylation, and intracellular transport of intestinal lactase-phlorizin hydrolase in rat. 311 97

A panel of monoclonal antibodies was produced against purified microvillus membranes of human small intestinal enterocytes. By means of these probes three disaccharidases (sucrase-isomaltase, lactase-phlorizin hydrolase, and maltase-glucoamylase) and four peptidases (aminopeptidase N, dipeptidylpeptidase IV, angiotension I-converting enzyme, and p-aminobenzoic acid peptide hydrolase) were successfully identified as individual entities by SDS PAGE and localized in the microvillus border of the enterocytes by immunofluorescence microscopy. The antibodies were used to study the expression of small intestinal hydrolases in the colonic adenocarcinoma cell line Caco 2. This cell line was found to express sucrase-isomaltase, lactase-phlorizin hydrolase, aminopeptidase N, and dipeptidylpeptidase IV, but not the other three enzymes. Pulse-chase studies with [35S]methionine and analysis by subunit-specific monoclonal antibodies revealed that sucrase-isomaltase was synthesized and persisted as a single-chain protein comprising both subunits. Similarly, lactase-phlorizin hydrolase was synthesized as a large precursor about twice the size of the lactase subunits found in the human intestine. Aminopeptidase N and dipeptidylpeptidase IV, known to be dimeric enzymes in most mammals, were synthesized as monomers. Transport from the rough endoplasmic reticulum to the trans-Golgi apparatus was considerably faster for the peptidases than for the disaccharidases, as probed by endoglycosidase H sensitivity. These results suggest that the major disaccharidases share a common biosynthetic mechanism that differs from that for peptidases. Furthermore, the data indicate that the transport of microvillus membrane proteins to and through the Golgi apparatus is a selective process that may be mediated by transport receptors.
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PMID:Expression and intracellular transport of microvillus membrane hydrolases in human intestinal epithelial cells. 389 50

The effects of an oral neomycin and penicillin regimen on intestinal bacteriology and on morphology and function of the small intestine of mice were investigated. Quantitative and qualitative stool cultures on selective media of the treated animals revealed only growth of yeast organisms. The treated animals developed enlargement of the ceca with fluid contents and watery stools, resembling characteristics of germfree animals. Radioautography with tritiated thymidine revealed an increased epithelial cell migration rate in the mice treated with the antibiotics for 3 to 5 wk. A slight increase in villus height was also noted. The treated male mice showed greater variance than the treated females in epithelial cell migration rates. Histochemical staining reactions showed a decrease in nonspecific esterase and in NADH dehydrogenase activity in the proximal gut of the antibiotic animals. Stains of distal gut and those for acid and alkaline phosphatase, NADPH dehydrogenase, lactic dehydrogenase, and succinic dehydrogenase were similar to the controls. A slight increase in sucrase activity and a slight decrease in lactase activity in the antibiotic animals was observed in contrast to control animals. Germfree mice, however, had greater sucrase and lactase activity. Transport of L-methionine was slightly reduced in the distal segment of the treated animals. Since the direction of these changes is away from the intestinal state observed in germfree animals, they are probably the result of the direct action of the antibiotics on the gut.
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PMID:Effects of neomycin and penicillin administration on mucosal proliferation of the mouse small intestine. With morphological and functional correlations. 438 18

The active site-directed inhibitor 4-nitrophenyl-beta-D-galactopyranosylmethyltriazene, previously shown (Fowler, A. V., Zabin, I., Sinnott, M. L., and Smith, P. J. (1978) J. Biol. Chem. 253, 5283-5285) to alkylate methionine 502 in lacZ beta-galactosidase, was used to label the second naturally occurring beta-galactosidase of Escherichia coli (ebgo). The reagent was also used to label two mutant forms of the enzyme (ebga and ebgb) selected for enhanced lactase activity. In the case of ebgo and ebga, 75 and 85% of the label, respectively, was incorporated into a tryptic peptide which is homologous (38% identity) to residues 483-503 of the lacZ beta-galactosidase sequence. In the ebgo and ebga enzymes, a serine probably is alkylated. In the case of the ebgb enzyme, 61% of the label is found on a tryptic peptide homologous (69% identity) with residues 457-468 of the lacZ beta-galactosidase. In this peptide, a glutamic acid and a tyrosine residue are both alkylated.
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PMID:The active site regions of lacZ and ebg beta-galactosidases are homologous. 641 10

The biosynthesis of pig small intestinal lactase-phlorizin hydrolase (EC 3.2.1.23-62) was studied by labelling of organ cultured mucosal explants with [35S]methionine. The earliest detactable form of the enzyme was an intracellular, membrane-bound polypeptide of Mr 225 000, sensitive to endo H as judged by its increased electrophoretic mobility (Mr 210 000 after treatment). The labelling of this form decreased during a chase of 120 min and instead two polypeptides of Mr 245 000 and 160 000 occurred, which both barely had their electrophoretic mobility changed by treatment with endo H. The Mr 160 000 polypeptide is of the same size as the mature lactase-phlorizin hydrolase and was the only form expressed in the microvillar membrane. Together, these data are indicative of an intracellular proteolytic cleavage during transport. The presence of leupeptin during labelling prevented the appearance of the Mr 160 000 form but not that of the Mr 245 000 polypeptide, suggesting that the proteolytic cleavage takes place after trimming and complex glycosylation. The proteolytic cleavage was not essential for the transport since the precursor was expressed in the microvillar membrane in the presence of leupeptin.
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PMID:Biosynthesis of intestinal microvillar proteins. Intracellular processing of lactase-phlorizin hydrolase. 643 Feb 96

Rats were made severely uremic with partial nephrectomy (24-hour creatinine clearance 10% of normal). Jejunal dipeptidase activities (substrates: glycyl-L-leucine, L-alanyl-L-proline, and L-methionyl-L-methionine), disaccharidase activities (maltase, sucrase, trehalase, and lactase) and morphology were studied. A highly significant increase in glycyl-L-leucine and L-methionyl-L-methionine dipeptidases was found in uremic rats compared with controls. Proline dipeptidase activities were unaltered. Disaccharidase activities showed a slight increase in sucrase in uremic rats; otherwise no change was found.
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PMID:Small intestinal dipeptidases and disaccharidases in experimental uremia in rats. 677 73

Explants of pig small intestine were maintained at 37 degrees C in organ culture for periods up to 24 h in a system using Trowell T-8 medium supplemented with 10% foetal-calf serum. The mucosal morphology was well preserved during culture, as judged by light and electron microscopy. The explant contents of protein and two brush-border enzymes, microvillus aminopeptidase (EC 3.4.11.2) and dipeptidyl peptidase IV (EC 3.4.14.5), were not significantly modified during culture compared with controls, but a moderate, continuous release of both protein and enzyme activities into the medium was observed. Continuous labelling with [35S]methionine resulted in an even incorporation of radioactivity in the protein components, and the rate of labelling only moderately decreased over the 24 h period. The polypeptide compositions of sucrase (EC 3.2.1.48)--isomaltase (EC 3.2.1.10), maltase--glucoamylase (EC 3.2.1.20) lactase (EC 3.2.1.23)--phlorizin hydrolase (EC 3.2.1.62), microvillus aminopeptidase and aspartate aminopeptidase (EC 3.4.11.7) synthesized during culture were studied, and some were found to be similar to those of the pro-forms of the enzymes isolated from animals that had had their pancreatic duct disconnected 3 days before being killed. These results confirmed earlier findings of the existence of pro-forms of some of the microvillar enzymes and thus indicate a low activity of pancreatic proteinases in the culture system.
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PMID:Biosynthesis of intestinal microvillar proteins. Characterization of intestinal explants in organ culture and evidence for the existence of pro-forms of the microvillar enzymes. 709 36

Human lactase-phlorizin hydrolase [EC 3.2.1.23-3.2.1.62] is a disaccharidase located in the microvillus membrane of small intestinal epithelial cells. The enzyme is synthesized as a precursor protein in the endoplasmic reticulum and in addition to being glycosylated is subsequently proteolytically processed to the mature microvillus membrane-bound form after passing the trans-Golgi compartment. We studied the oligomerization of human lactase-phlorizin hydrolase in transfected polarized Madin Darby canine kidney cells using metabolic labeling and sucrose-density centrifugation analysis. We detected high mannose dimers of the lactase-phlorizin hydrolase precursor molecule after metabolic labeling with [35S]methionine at 37 and 15 degrees C. In addition, both complex-glycosylated lactase-phlorizin hydrolase precursor molecule and the mature microvillus membrane-bound enzyme showed this oligomeric structure. Chemical crosslinking resulted in the detection of covalently crosslinked lactase-phlorizin hydrolase dimers after sodium dodecyl sulfate polyacrylamide gel electrophoresis. These results provide evidence that oligomerization of lactase-phlorizin hydrolase is an early event and begins in the endoplasmic reticulum.
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PMID:Human lactase-phlorizin hydrolase: evidence of dimerization in the endoplasmic reticulum. 748

We have previously shown that fetal exposure to ethanol in rats produces both structural and biochemical abnormalities in absorptive enterocytes. Among the indicators of injury are derangements in the expression of lactase-phlorizin hydrolase (LPH), which is an essential enzyme for the assimilation of milk. In an animal model of fetal alcohol syndrome, unsuckled newborn rats prenatally exposed to maternal ethanol revealed a 10- to 15-fold increase in the number of LPH mRNA molecules per absorptive enterocyte, compared with controls (Estrada et al., Alcohol. Clin. Exp. Res. 20:1662-1668, 1996). However, lactase activity per cell was similar in both groups. The aim of this study was to characterize the effect of prenatal exposure to ethanol on the processing of LPH mRNA and protein. RNase protection assays using 3'- and 5'-directed antisense RNA probes revealed that the LPH mRNA from ethanol-exposed pups is full length. However, metabolic labeling, followed by immunoprecipitation using an anti-LPH monoclonal antibody, demonstrated a significant alteration in LPH protein processing. Intestinal explants from 21-day ethanol-exposed fetuses that were chased 30 min after a [35S]methionine pulse showed greater amounts of newly synthesized LPH precursors (205 and 220 kDa) and low molecular weight degradation products than controls. However, despite the increases in LPH precursor, the amount of 130 kDa mature LPH was similar in ethanol-exposed and control explants. These data suggest an increase in intracellular degradation of LPH precursor in rats prenatally exposed to ethanol, which occurs before its insertion into the microvillus membrane. Biosynthesis of LPH appears to be upregulated at the transcriptional level, which overcomes the degradation of LPH precursor during processing.
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PMID:Defective intracellular processing of lactase-phlorizin hydrolase protein in rats prenatally exposed to ethanol. 972 93

Glucocorticoids from endogenous and exogenous sources accelerate maturation of brush-border membrane (BBM) hydrolases in omnivorous laboratory rodents and pigs. Less is known for carnivores, and whether the route of administration (oral or systemic) has an influence. The present study examined the influence of administering cortisol (hydrocortisone succinate, 5 mg/kg-day) to mink during postnatal week 4, just prior to weaning, on small intestine glucose and amino acid (aspartate, leucine, lysine, methionine, proline) absorption and on the activities of BBM disaccharidases and peptidases. Kits treated with cortisol were smaller (P<0.05), but had small intestines that were proportionally larger (P<0.05 for length and mass per kg body weight, but not for mucosal mass) than control kits with higher rates of absorption for most nutrients, except leucine, and increased activities of most BBM hydrolases, except lactase. As a consequence, cortisol increased hydrolytic and absorptive capacities of the entire small intestine, with the responses more pronounced when the cortisol was given orally. These findings indicate administration of cortisol stimulates growth of the developing mink small intestine, but does not accelerate the postnatal declines in nutrient transport, and may be a dam-to-kit signal that prepares suckling mink to digest and absorb the adult diet.
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PMID:Cortisol increases the activities of intestinal apical membrane hydrolases and nutrient transporters before weaning in mink (Mustela vison). 1634 90


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