Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.2.1.23 (beta-galactosidase)
14,648 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Microbial-derived beta-galactosidase (beta-gal) enzyme preparations improve in vivo lactose digestion and tolerance through enhanced gastrointestinal digestion of lactose. Three different beta-gal preparations, Lactogest (soft gel capsule), Lactaid (caplet), and DairyEase (chewable tablet) and placebo were fed to lactose maldigesters with either 20 g or 50 g of lactose to compare the efficacy of these products and to further establish a dose-response relationship for use. All enzyme preparations dramatically reduced both the peak and total breath hydrogen production when fed with milk containing 20 g of lactose. Four capsules of Lactogest, two caplets of Lactaid, or two tablets of DairyEase (each treatment containing approx 6000 IU) reduced total hydrogen production significantly (P < 0.05) below that observed with two capsules of Lactogest (containing approx 3000 IU) in a stoichiometric manner. Symptoms were significantly (P < 0.05) less severe with all the beta-gal products. In contrast, with 50 g of lactose in water, peak and total hydrogen production was modestly, but not significantly reduced by the enzyme treatment. Furthermore, symptom scores for bloating, cramping, nausea, pain, diarrhea, and flatus were not different between treatments and the control. The 50-g lactose dose appeared to overwhelm the ability of either 3000 or 6000 IU of beta-gal to assist significantly with lactose digestion. Results from these studies demonstrate the relative equivalency of chewable, caplet, and soft-gel beta-gal products, based on IUs of enzyme fed.
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PMID:Comparative effects of exogenous lactase (beta-galactosidase) preparations on in vivo lactose digestion. 822 76

Twelve healthy volunteers were studied for two test periods, at the beginning of which they ingested a diarrheogenic load (60 g) of lactulose in 350 mL water with 10 g polyethylene glycol 4000 (PEG); the two periods were separated by a lactulose feeding period of 8 d, during which a nondiarrheogenic load (20 g) of lactulose was taken twice daily. The transit time and flow rates of water and lactulose in the distal ileum of four subjects were not different before and after the lactulose feeding period. In the other eight subjects, stool weight and frequency, fecal pH, and fecal outputs of carbohydrates and osmotic moieties after the ingestion of 60 g lactulose dropped significantly (P < 0.05) after the lactulose feeding period, whereas the orofecal transit time and fecal concentrations of beta-galactosidase and lactic acid increased (P < 0.05). We conclude that changes in colonic function induced by prolonged exposure to a nondiarrheogenic amount of lactulose mitigate the severity of the diarrhea because of the larger dose of lactulose.
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PMID:Can diarrhea induced by lactulose be reduced by prolonged ingestion of lactulose? 823 48

The effect of salts (KI, KBr, NaCl, KCl, KF, phosphate, and Na2SO4) on the stability of beta-galactosidase in aqueous solution was studied from the aspect of changes in water mobility. At salt concentrations up to 200 mM, the inactivation rate of beta-galactosidase in all the salt solutions studied increased with increasing salt concentration. At higher concentrations, those salts which had little effect on the spin-lattice relaxation time, T1, of water (KI, KBr, and KCl) continued to increase the inactivation rate of beta-galactosidase with increasing concentration, while those salts which decreased the T1 of water (KF, phosphate, and Na2SO4) decreased the inactivation rate. It appeared that the decrease in water mobility caused by KF, phosphate, and Na2SO4 resulted in stabilization of beta-galactosidase. The results indicate that water mobility is an important factor in the denaturation rate of proteins.
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PMID:The effect of salts on the stability of beta-galactosidase in aqueous solution, as related to the water mobility. 827 11

Cascade polymers also known as Starburst dendrimers are spheroidal polycations that can be synthesized with a well-defined diameter and a precise number of terminal amines per dendrimer. We show, using luciferase and beta-galactosidase containing plasmids, that dendrimers mediate high efficiency transfection of a variety of suspension and adherent cultured mammalian cells. Dendrimer-mediated transfection is a function both of the dendrimer/DNA ratio and the diameter of the dendrimer. Maximal transfection of luciferase are obtained using a diameter of 68 A and a dendrimer to DNA charge ratio of 6/1 (terminal amine to phosphate). Expression is unaffected by lysomotrophic agents such as chloroquine and only modestly affected (2-fold decrease) by the presence of 10% serum in the medium. Cell viability, as assessed by dye reduction assays, decreases by only 30% at 150 micrograms dendrimer/mL in the absence of DNA and about 75% in the presence of DNA. Under similar conditions polylysine causes a complete loss of viability. Gene expression decreased by 3 orders of magnitude when the charge ratio is reduced to 1:1. When GALA, a water soluble, membrane-destabilizing peptide, is covalently attached to the dendrimer via a disulfide linkage, transfection efficiency of the 1:1 complex is increased by 2-3 orders of magnitude. The high transfection efficiency of the dendrimers may not only be due to their diameter and shape but may also be caused by the pKa's (3.9 and 6.9) of the amines in the polymer. The low pKa's permit the dendrimer to buffer the pH change in the endosomal compartment. The characteristics of precise control of structure, favorable pKa's, and low toxicity make the dendrimers suitable for gene-transfer vehicles.
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PMID:Polyamidoamine cascade polymers mediate efficient transfection of cells in culture. 827 23

A new membrane filter agar medium (MI agar) containing a chromogen, indoxyl-beta-D-glucuronide, and a fluorogen, 4-methylumbelliferyl-beta-D-galactopyranoside, was developed to simultaneously detect and enumerate Escherichia coli and total coliforms (TC) in water samples on the basis of their enzyme activities. TC produced beta-galactosidase, which cleaved 4-methylumbelliferyl-beta-D-galactopyranoside to form 4-methylumbelliferone, a compound that fluoresced under longwave UV light (366 nm), while E. coli produced beta-glucuronidase, which cleaved indoxyl-beta-D-glucuronide to form a blue color. The new medium TC and E. coli recoveries were compared with those of mEndo agar and two E. coli media, mTEC agar and nutrient agar supplemented with 4-methylumbelliferyl-beta-D-glucuronide, using natural water samples and spiked drinking water samples. On average, the new medium recovered 1.8 times as many TC as mEndo agar, with greatly reduced background counts (< or = 7%). These differences were statistically significant (significance level, 0.05). Although the overall analysis revealed no statistically significant difference between the E. coli recoveries on MI agar and mTEC agar, the new medium recovered more E. coli in 16 of 23 samples (69.6%). Both MI agar and mTEC agar recovered significantly more E. coli than nutrient agar supplemented with 4-methylumbelliferyl-beta-D-glucuronide. Specificities for E. coli, TC, and noncoliforms on MI agar were 95.7% (66 of 69 samples), 93.1% (161 of 173 samples), and 93.8% (61 of 65 samples), respectively. The E. coli false-positive and false-negative rates were both 4.3%. This selective and specific medium, which employs familiar membrane filter technology [corrected] to analyze several types of water samples, is less expensive than the liquid chromogen and fluorogen media and may be useful for compliance monitoring of drinking water.
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PMID:New medium for the simultaneous detection of total coliforms and Escherichia coli in water. 828 60

The application of cationic liposome reagents has advanced DNA and mRNA transfection research in vitro, and data are accumulating which show their utility for in vivo gene transfer. However, chemical structure-activity data leading to a better mechanistic understanding of their biological activity is still limited. Most of the cationic lipid reagents in use today for this application are formulated as liposomes containing two lipid species, a cationic amphiphile and a neutral phospholipid, typically dioleoylphosphatidylethanolamine (DOPE). The studies reported here examine the effects of some systematic chemical structural changes in both of these lipid components. Cationic and neutral phospholipids were formulated together as large multilamellar vesicles (MLV) or small sonicated unilamellar vesicles (SUV) in water, and each formulation was assayed quantitatively in 96-well microtiter plates under 64 different assay conditions using COS.7 cells and an RSV-beta-galactosidase expression plasmid. The cationic lipid molecules used for these studies were derived from a novel series of 2,3-dialkyloxypropyl quaternary ammonium compounds containing a hydroxyalkyl moiety on the quaternary amine. A homologous series of dioleylalkyl (C18:1) compounds containing increasing hydroxyalkyl chain lengths on the quaternary amine were synthesized, formulated with 50 mol % DOPE, and assayed for transfection activity. The order of efficacy was ethyl > propyl > butyl > pentyl > 2,3-dioleyloxypropyl-1-trimethyl ammonium bromide (DOTMA). DOTMA, which is commercially available under the trademark Lipofectin Reagent, lacks a hydroxyalkyl moiety on the quaternary amine. A homologous series of hydroxyethyl quaternary ammonium derivatives with different alkyl chain substitutions were synthesized, formulated with 50 mol % DOPE, and assayed in the transfection assay. The order of transfection efficacy was dimyristyl (di-C14:0) > dioleyl (di-C18:1) > dipalmityl (di-C16:0) > disteryl (di-C18:0). The addition of 100 microM chloroquine in the transfection experiment enhanced the activity of the dioleyl compound by 4-fold and decreased the activity of the dimyristyl compound by 70%. For each of the compounds and formulations examined in this report, large multilamellar vesicles (MLV; diameter 300-700 nm) were more active than small unilamellar vesicles (SUV; diameter 50-100 nm). The neutral phospholipid requirements for transfection activity in COS.7 cells with these cationic lipid molecules were examined.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Enhanced gene delivery and mechanism studies with a novel series of cationic lipid formulations. 830 May 83

Aggregates formed during storage of freeze-dried beta-galactosidase were compared with those formed in solutions. Freeze-dried beta-galactosidase aggregated during storage in the presence of moisture, producing a protein precipitate which was soluble in guanidine hydrochloride solution but not in buffer solution. The protein precipitate dissolved in guanidine solution exhibited a large molecular size by high-performance size exclusion chromatography and converted to proteins of original size in the presence of dithiothreitol. It is suggested that the aggregation involves chemical interaction via covalent disulfide bonding. In contrast, beta-galactosidase in aqueous solution aggregated without formation of protein precipitates. Soluble aggregates were converted to proteins of original size in guanidine solution without dithiothreitol, suggesting noncovalent bonding. The difference in aggregation behavior may be ascribed to the difference in the water:protein ratio. We propose that inactivation of beta-galactosidase is due to formation of thermally denatured (unfolded) protein, which aggregates dependent on the water:protein ratio, either via noncovalent interactions at a high water:protein ratio in solution or via covalent interaction at a low water:protein ratio in the freeze-dried state.
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PMID:Aggregates formed during storage of beta-galactosidase in solution and in the freeze-dried state. 832 32

The inactivation of freeze-dried beta-galactosidase during storage was studied, focusing on the effect of water mobility as measured by the spin-lattice relaxation time, T1, of water using 17O NMR. Inactivation of beta-galactosidase lyophilized from phosphate buffer solution was studied as a function of water content, which in turn affected the T1 of water. An increase in the water content of freeze-dried beta-galactosidase brought about an increase in the T1 of water, as well as a rise in pH. For the freeze-dried enzyme with sufficient water content to be dissolved, the inactivation rate was related to the T1 of water rather than to the pH change. It is suggested that as the water content increases, the mobility of water around the enzyme increases, resulting in enhanced enzyme inactivation. The freeze-dried samples with limited moisture showed inactivation rates faster than those expected from the pH and water mobility, suggesting that the inactivation mechanism is different from that for the freeze-dried enzyme with a larger amount of water. Inactivation of beta-galactosidase in solutions was also studied as a function of phosphate buffer and sodium chloride concentrations, which in turn affected the T1 of water. Because the inactivation rate increased with increasing salt concentrations and the rate extrapolated to zero concentration was negligible, inactivation of the freeze-dried enzyme was apparently induced by the salts used as additives for lyophilization. The enhancing effect of phosphate buffer components, however, was reduced at higher concentrations, an effect related to the decrease in the T1 of water. This result may be ascribed to the decrease in water mobility caused by phosphate buffer components and is consistent with the observation that the inactivation rate of the freeze-dried enzyme with a relatively large amount of water decreased with decreasing T1 of water.
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PMID:Stability of beta-galactosidase, a model protein drug, is related to water mobility as measured by 17O nuclear magnetic resonance (NMR). 843 45

We developed a chemiluminometric assay of beta-galactosidase in coliform bacteria, using a phenylgalactose-substituted 1,2-dioxetane derivative as a substrate. Permeabilization of cells is required to ensure the efficient cellular uptake of this compound. By this method, one coliform seeded in 100 ml of sterile water can be detected after a 6- to 9-h propagation phase followed by a 45-min enzyme assay in the presence of polymyxin B. Compared with fluorometry and colorimetry, chemiluminometry afforded 4- and 1,000-fold increases in sensitivity and 1- and 6-h increases in the speed of detection, respectively.
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PMID:Development of a sensitive chemiluminometric assay for the detection of beta-galactosidase in permeabilized coliform bacteria and comparison with fluorometry and colorimetry. 853 20

The kinetics of hydrolysis of a series of synthetic substrates by two experimentally evolved forms ('evolvants'), ebgabcd and ebgabcde, of the second beta-galactosidase of Escherichia coli have been measured. The ebgabcd enzyme differs from the wild-type (ebgo) enzyme by Asp92-->Asn (a) and Trp977-->Cys (b) changes in the large subunit, as well as two changes hitherto considered to have no kinetic effect, Ser979-->Gly in the large subunit (c) and Glu122-->Gly in the small subunit (d). The enzyme ebgabcde contains in addition a Glu93-->Lys change in the large subunit (e). Comparison of ebgabcd with ebgab [Elliott, K, Sinnott, Smith, Bommuswamy, Guo, Hall and Zhang (1992) Biochem. J. 282, 155-164] indicates that the c and d changes in fact accelerate the hydrolysis of the glycosyl-enzyme intermediate by a factor of 2.5, and also decrease the charge on the aglycone oxygen atom at the first transition state; the charge on the glycone, however, is unaltered [see K, Konstantinidis, Sinnott and Hall (1993) Biochem. J. 291, 15-17]. The e mutation causes a fall in the degalactosylation rate of about a factor of 3, and its occurrence only together with c and d mutations [Hall, Betts and Wootton (1989) Genetics 123, 635-648] suggests that degalactosylation of a hypothetical ebgabe enzyme would be so slow that the enzyme would have no biological advantage over the ancestral ebgab. The transfer products from galactosyl-ebgabcd and galactosyl-ebgabcde to high concentrations to glucose have been measured; the predominant product is allolactose, but significant quantities of lactose are also formed; however, at apparent kinetic saturation of the galactosyl-enzyme, hydrolysis rather than transfer is the preponderant pathway. A knowledge of the rates of enzyme-catalysed exchange of 18O from [1-18O]galactose to water permits the construction of the free-energy profiles for hydrolysis of lactose by begabcd and ebgabcde. As with the other evolvants, changes in the profile away from the rate-determining transition state are essentially random, and there is no correlation between the changes in the free energies of intermediates and of their flanking transition states. We consider the aggregate of our kinetic data on the ebg system to be telling experimental support for the theoretical objections of Pettersson [Pettersson (1992) Eur. J. Biochem. 206, 289-295 and previous papers] to the Albery-Knowles theory of the evolution of enzyme kinetic activity.
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PMID:Catalytic consequences of experimental evolution: catalysis by a 'third-generation' evolvant of the second beta-galactosidase of Escherichia coli, ebgabcde, and by ebgabcd, a 'second-generation' evolvant containing two supposedly 'kinetically silent' mutations. 855 46


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