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)

Sucrase-isomaltase (SI), trehalase (T) and lactase-beta-glucosidase (LG) activities were assessed histochemically in samples of colorectal adenomas (11 tubular, 12 tubulovillous, 10 villous) and 30 adenocarcinomas obtained by biopsy during colonoscopy or from specimens removed by surgical intervention. Small samples of tumor tissue, tissue of the transitional zone and of macroscopically normal mucosa were quenched in heptan cooled in an acetone-dry ice mixture. Cryostat sections, transferred to non-precooled slides and in some cases to semipermeable membranes, were dried and subjected to the histochemical reactions for SI, T and LG. Sucrose, 2-naphthyl, 6-Br-2-naphthyl, and 5-Br-4-Cl-3-indoxyl alpha-D-glucosides, trehalose, and 5-Br-4-Cl-3-indoxyl-beta-D-fucoside were used as substrates. Sections of jejunal biopsies with normal activities of brush border glycosidases were used as controls. From samples of 5 adenomas, 5 adenocarcinomas and collected rests of jejunal biopsies with a normal finding 10% (w/vol) homogenates in 2% Triton X-100 were prepared. Homogenates were frozen and thawed 3 times and their supernatants subjected to isoelectric focusing on polyacrylamide gel plates. Zymograms were developed with the same methods as for the detection of alpha-glucosidases in sections. In no colorectal tumor LG was present. SI was found in 70% adenocarcinomas, 50% villous, 25% tubulovillous and 19% tubular adenomas when the method with sucrose, glucose oxidase-peroxidase and 3,3'-diaminobenzidine was used. Hardly discernible traces of activity were found in tumors with azo-coupling reactions applied at pH 5, 6 and 6.5. No reaction was detected with the indigogenic method applied at pH above 6.0. However, jejunal biopsies displayed very strong reactions confined to the brush border of enterocytes under the same conditions. A strongly positive reaction was seen in 7 of 12 tumors investigated recently when the indigogenic reaction was applied at pH below 6.0 (particularly at pH 5.0). In this case the deposition of indigo was due to membrane and lysosomal alpha-glucosidases of the tumor cells and lysosomal alpha-glucosidase of macrophages and leukocytes. These findings were corroborated by zymograms. T was detected in the same tumors as SI; its activity was lower, however. SI activity in colorectal tumors is a useful, but not general marker of these tumors.
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PMID:Sucrase-isomaltase and other brush border glycosidases in colorectal tumors. 886 57

The cytosolic beta-glucosidase (EC 3.2.1.21) present in the livers of mammalian species is distinguished by its broad specificity for sugars and its preference for hydrophobic aglycones. We purified the cytosolic beta-glucosidase from guinea pig liver and sequenced 142 amino acid residues contained within 12 trypsin digest fragments. Using degenerate oligonucleotide primers deduced from the peptide sequences, a 622 bp cytosolic beta-glucosidase cDNA was amplified by reverse-transcriptase PCR, using total guinea pig liver RNA as template. The 'rapid amplification of cDNA ends (RACE)' method [Frohman (1993) Methods Enzymol. 218, 340-356] was used to synthesize the remaining segments of the full-length cDNA. The complete cDNA contained 1671 nucleotides with an open reading frame coding for 469 amino acid residues. The amino acid sequence deduced from the cDNA sequence included the amino acid sequences of all 12 trypsin digest fragments derived from the purified enzyme. Amino acid sequence analysis indicates that the guinea pig liver cytosolic beta-glucosidase is a Family 1 beta-glycosidase and that it is most closely related to mammalian lactase-phlorizin hydrolase. These results suggest that the cytosolic beta-glucosidase and lactase-phlorizin hydrolase diverged from a common evolutionary precursor.
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PMID:Primary structure of the cytosolic beta-glucosidase of guinea pig liver. 892 Sep 87

Aqueous ethanol extracts from the immature fruits and stalks of bluebell (Hyacinthoides non-scripta) were subjected to various ion-exchange column chromatographic steps to give 1,4-dideoxy-1,4-imino-D-arabinitol (1),2(R),5(R)-bis(hydroxymethyl)-3(R),4(R)-dihydroxypyrrolidine (DMDP) (2), 6-deoxy-6-C-(2,5-dihydroxyhexyl)-DMDP (3),2,5-dideoxy-2,5-imino-DL-glycero-D-manno-heptitol (homoDMDP)(4),homoDMDP-7-O-apioside (5), homoDMDP-7-O-beta-D-xylopyranoside (6), (1S*,2R*,3R*,5R*,7aR*)-1,2-dihydroxy-3,5- dihydroxymethylpyrrolizidine (7), and (1S*,2R*,3R*,5R*,6R*,7R*,7aR*)-3-hydroxymethyl-5-methyl-1,2,6,7 tetrahydroxypyrrolizidine (8). Bulbs of Scilla campanulata (Hyacinthaceae) yielded (1S*,2R*,3R*,5S*,7aR*)-1,2-dihydroxy-3,5-dihydroxy-methylpyrrol izidine (9) in addition to compounds 1-7. Compounds 3,6,7,8, and 9 are new natural products. Compound 4 is a potent competitive inhibitor with K(i) values of 1.5 microM for Caldocellum saccharolyticum beta-glucosidase and 2.2 microM for bovine liver beta-galactosidase. The 7-O-beta-D xyloside 6 was a stronger competitive inhibitor than 4 of C saccharolyticum beta-glucosidase and rat intestinal lactase, with K(i) values of 0.06 and 0.07 microM, respectively, but a weaker inhibitor of bovine liver beta-galactosidase. Furthermore, compound 4 is also a competitive inhibitor (K(i) = 1.8 microM) of porcine kidney trehalase, but 6 was inactive against this enzyme.
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PMID:Polyhydroxylated pyrrolidine and pyrrolizidine alkaloids from Hyancinthoides non-scripta and Scilla campanulata. 1051 98

Two beta-glycosidases (BG) (Mr 47,000 and Mr 50,000) were purified from Spodoptera frugiperda (Lepidoptera: Noctuidae) midguts. These two polypeptides associate or dissociate depending on the medium ionic strength. The Mr 47,000 BG probably has two active sites. One of the putative active sites (cellobiase site) hydrolyses p-nitrophenyl beta-D-glucoside (NPbetaGlu) (79% of the total activity in saturated enzyme), cellobiose, amygdalin and probably also cellotriose, cellotetraose and cellopentaose. The cellobiase site has four subsites for glucose residue binding, as can be deduced from cellodextrin cleavage data. The enzymatic activity in this site is abolished after carbodiimide modification at pH 6.0. Since the inactivation is reduced in the presence of cellobiose, the results suggest the presence of a carboxylate as a catalytic group. The other active site of Mr 47,000 BG (galactosidase site) hydrolyses p-nitrophenyl beta-D-galactoside (NPbetaGal) better than NPbetaGlu, cleaves glucosylceramide and lactose and is unable to act on cellobiose, cellodextrins and amygdalin. This active site is not modified by carbodiimide at pH 6.0. The Mr 47,000 BG N-terminal sequence has high identity to plant beta-glycosidases and to mammalian lactase-phlorizin hydrolase, and contains the QIEGA motif, characteristic of the family of glycosyl hydrolases. The putative physiological role of this enzyme is the digestion of glycolipids (galactosidase site) and di- and oligosaccharides (cellobiase site) derived from hemicelluloses, thus resembling mammalian lactase-phlorizin hydrolase.
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PMID:Purification and properties of a beta-glycosidase purified from midgut cells of Spodoptera frugiperda (Lepidoptera) larvae. 1104 60

Cytosolic beta-glucosidase (EC 3.2.1.21) from mammalian liver is a member of the family 1 glycoside hydrolases and is known for its ability to hydrolyse a range of beta-D-glycosides, including beta-D-glucoside and beta-D-galactoside. We therefore refer to this enzyme as cytosolic beta-glycosidase. We cloned the cDNA encoding the human cytosolic beta-glycosidase by performing PCR on cDNA prepared from total human liver RNA. Specific primers were based on human expressed sequence tags found in the expressed sequence tag database. The cloned cDNA contained 1407 nt with an open reading frame encoding 469 amino acid residues. Amino acid sequence analysis indicates that human cytosolic beta-glycosidase is most closely related to lactase phlorizin hydrolase and klotho protein. The enzyme was characterized by using cell lysates of COS-7 cells transfected with a eukaryotic expression vector containing the cDNA. The biochemical, kinetic and inhibition properties of the cloned enzyme were found to be identical with those reported for the enzyme purified from human liver.
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PMID:Cloning and characterization of human liver cytosolic beta-glycosidase. 1138 1

An obligatory step in the mammalian nutritional utilization of pyridoxine-5'-beta-D-glucoside (PNG) is the intestinal hydrolysis of its beta-glucosidic bond that releases pyridoxine (PN). This laboratory previously reported the purification and partial characterization of a novel cytosolic enzyme, designated PNG hydrolase, which hydrolyzed PNG. An investigation of the subcellular distribution of intestinal PNG hydrolysis found substantial hydrolytic activity in the total membrane fraction, of which 40-50% was localized to brush border membrane. To investigate the possible role of a brush border beta-glucosidase in the hydrolysis of PNG, lactase phlorizin hydrolase (LPH) was purified from rat small intestinal mucosa. LPH hydrolyzed PNG with a K(m) of 1.0 +/- 0.1 mm, a V(max) of 0.11 +/- 0.01 micromol/min.mg protein, and a k(cat) of 1.0 s(-1). LPH-catalyzed PNG hydrolysis was inhibited by glucose, lactose, and cellobiose but not by PN. Specific blockage of the phlorizin hydrolase site of LPH using 2',4'-dintrophenyl-2-fluoro-2-deoxy-beta-D-glucopyranoside did not reduce PNG hydrolysis. Evidence of transferase activity was also obtained. Reaction mixtures containing LPH, PNG, and lactose yielded the formation of another PN derivative that was identified as a pyridoxine disaccharide. These results indicate that LPH may play an important role in the bioavailability of PNG, but further characterization is needed to assess its physiological function.
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PMID:Enzymatic hydrolysis of pyridoxine-5'-beta-D-glucoside is catalyzed by intestinal lactase-phlorizin hydrolase. 1202 80

Pyridoxine-5'-beta-D-glucoside (PNG) is a major form of vitamin B-6 in plant foods that exhibits partial bioavailability as vitamin B-6 in humans. We previously identified an intestinal mucosal cytosolic PNG hydrolase that catalyzes the partial hydrolysis of PNG absorbed without prior deglycosylation. Recent observations that the brush border membrane also catalyzes PNG hydrolysis led to the hypothesis that PNG hydrolysis may be another function of the beta-glucosidase lactase-phlorizin hydrolase (LPH) and, thus, brush border PNG hydrolysis would undergo a developmental decline similar to that of lactose hydrolysis. In this study, the relationships among hydrolytic activities in small intestinal cytosolic and brush border fractions in rats (n = 9 per group) of various ages (1-2 d and 2, 4, 8, 12 and 24 wk) were examined. In vitro specific activities toward PNG and lactose were greater in brush border than cytosol, and these were greater in newborn rats than in all other age groups (P < 0.01). Brush border activities toward PNG and lactose and were closely correlated (r = 0.84; P < 0.0001). These findings suggest that the hydrolysis of PNG is catalyzed at least partially at the brush border and that the bioavailability of PNG may be influenced by the residual LPH activity in children and adults.
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PMID:Intestinal brush border membrane catalyzes hydrolysis of pyridoxine-5'-beta-D-glucoside and exhibits parallel developmental changes of hydrolytic activities toward pyridoxine-5'-beta-D-glucoside and lactose in rats. 1222 Dec 31

We have previously identified and purified a novel beta-glucosidase, designated PNGH (pyridoxine-5'-beta-D-glucoside hydrolase), from the cytosolic fraction of pig intestinal mucosal. PNGH catalyses the hydrolysis of PNG (pyridoxine-5'-beta-D-glucoside), a plant derivative of vitamin B6 that exhibits partial nutritional bioavailability in humans and animals. Preliminary amino acid sequence analysis indicated regions of close similarity of PNGH to the precursor form of LPH (lactase-phlorizin hydrolase), the beta-glucosidase localized to the brush-border membrane. We report in the present study amino acid sequence data for PNGH and results of Northern blot analyses, upon which we propose a common genomic origin of PNGH and LPH. Internal Edman sequencing of the PNGH band isolated by SDS/PAGE yielded data for 16 peptides, averaging 10.8 amino acids in length. These peptides from PNGH (approx. 140 kDa) were highly similar to sequences existing over most of the length of the >200 kDa precursor of rabbit LPH; however, we found no PNGH sequences that corresponded to approx. 350 amino acids between positions 463 and 812 of the LPH precursor, a region encoded by exon 7 of the LPH precursor gene (amino acids 568-784), and no sequences that corresponded to regions near the N-terminus. MS analysis of tryptic peptides yielded 25 peptides, averaging 15 amino acids, with masses that matched segments of the rabbit LPH precursor. Northern blot analysis of pig and human small intestinal polyadenylated mRNA using a non-specific LPH cDNA probe showed an expected approx. 6 kb transcript of the LPH precursor, but also an approx. 4 kb transcript that was consistent with the size predicted from the PNGH protein mass. Using a probe specific to the region encoded by exon 7, hybridization occurred only with the 6 kb transcript. Based on these observations, we propose that both PNGH and LPH enzymes have the same genomic origin, but differ in transcriptional and, possibly, post-translational processing.
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PMID:Partial amino acid sequence and mRNA analysis of cytosolic pyridoxine-beta-D-glucoside hydrolase from porcine intestinal mucosa: proposed derivation from the lactase-phlorizin hydrolase gene. 1497 28

The present study analyzed the existence of carbohydrases in camel pancreas compared to some other ruminants. Disaccharidases (maltase, cellobiase, lactase, trehalase and sucrase), glucoamylase and alpha-amylase were detected in pancreas of camel, sheep, cow and buffalo. Enzyme levels in sheep were lower than in the other ruminants. The highest level was detected for alpha-amylase (EC 3.2.1.2). Moderate activity levels were detected for glucoamylase (EC 3.2.1.3) and maltase (EC 3.2.1.20), while other disaccharidases showed very low activity. The results suggested that, in addition to alpha-amylase, glucoamylase and maltase may be synthesized and secreted from pancreas to the small intestine in ruminants. Camel pancreatic glucoamylase was purified and characterized. The purification procedure included glycogen precipitation and chromatography on DEAE-Sepharose and Sepharose 6B. The molecular mass was 58 kDa for native and denatured enzyme using gel filtration and SDS-PAGE, respectively. The enzyme had a pH optimum at 5.5 and a Km of 10 mg starch/mL with more affinity toward potato soluble starch than the other carbohydrates. Glucoamylase had a temperature optimum at 50 degrees C with heat stability up to 30 degrees C. The effect of different cations and inhibitors was examined. The camel pancreatic glucoamylase may possess an essential thiol.
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PMID:Carbohydrases in camel (Camelus dromedarius) pancreas. Purification and characterization of glucoamylase. 1562 12

Glycosidase activity influences the intestinal absorption of glycosides. Our previous study in rats suggested that disaccharide conjugates might be prototypes for pre-prodrugs aiming at the Na(+)/glucose co-transporter-mediated transport of prodrugs (drug glucoside) as a novel absorption pathway. One of the crucial factors is the formation of a glucoside drug from the disaccharide conjugate. Since there is a large species difference in metabolism, it is necessary to examine the cells and/or enzymes derived from human tissue to confirm this concept. In this paper, we kinetically characterized the glycosidase activity of disaccharide conjugates in Caco-2 cells. Disaccharide conjugates of p-nitrophenol (p-NP) (p-NP beta-cellobioside, p-NP beta-lactoside and p-NP beta-maltoside) were hydrolysed to p-NP beta-glucoside. beta-glucosidase or beta-galactosidase (lactase/phloridzin hydrolase, LPH) and alpha-glucosidase (sucrase-isomaltase) had different pH-dependent activities for disaccharide conjugates. At neutral pH, LPH has low affinity and low capacity, and sucrase-isomaltase has high affinity and high capacity, whereas at acid pH, LPH has high affinity and low capacity, and sucrase-isomaltase has low affinity and high capacity. The hydrolysis clearance calculated with Vmax/Km indicated that sucrase-isomaltase activity is much higher than LPH activity at either neutral or acid pH in Caco-2 cells. Since the hydrolysis rate of the disaccharide conjugate was highly dependent on the pH value and type of glycoside linkage, the appropriate selection of a glycoside form after consideration of these differences is the key to designing a sugar-conjugate prodrug.
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PMID:Kinetic characterization of glycosidase activity from disaccharide conjugate to monosaccharide conjugate in Caco-2 cells. 1590 56


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