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)

4-Methylumbelliferyl (4-MU) glycosides of N-acetylglucosamine oligosaccharides were used as substrates to detect expression of a Streptomyces chitinase in Escherichia coli. Low levels of enzyme were detected when S. plicatus DNA was cloned into a bacteriophage lambda vector (EMBL-4). Subcloning into E. coli plasmids also gave low but detectable levels of enzyme expression. High level expression was achieved by resection of the cloned S. plicatus DNA with Bal31 followed by in-frame fusion to the amino-terminal peptide sequence of beta-galactosidase found in the pUC vectors. The Streptomyces chitinase was secreted into the periplasmic space of E. coli, and its signal sequence was removed. We characterized the activity of the cloned enzyme and compared it to three other purified Streptomyces plicatus chitinases with respect to hydrolysis of the 4-MU oligosaccharides. We found that two of the enzymes form 4-methylumbelliferone much more rapidly from the 4-MU disaccharide than from the trisaccharide. These same enzymes convert the 4-MU trisaccharide primarily to diacetylchitobiose and the 4-MU monosaccharide, a nonfluorescent product. The latter compound is not hydrolyzed appreciably by any of the enzymes. On the basis of these results, we suggest a new definition of "exo" and "endo" chitinase that differs from that found in the literature. We propose that exochitinase activity be defined as processive action starting at the nonreducing ends of chitin chains with release of successive diacetylchitobiose units, and that endochitinase activity be defined as random cleavage at internal points in chitin chains.
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PMID:Cloning and expression of a Streptomyces plicatus chitinase (chitinase-63) in Escherichia coli. 327 46

Recently we have describe a simple efficient chemical method of generating an asparagine side-chain linker with beta-stereochemistry at the anomeric position of neutral oligosaccharides. We now report the 1-N-glycyl beta-derivatization of sialylated saccharides. Several neoglycoconjugates formed using these N-linked inter-mediates were investigated for their usefulness in probing carbohydrate-protein interactions. First, biotinyl derivatives of two xylose/fucose class plant-type oligosaccharides purified from horseradish peroxidase were effective in demonstrating the carbohydrate specificity of polyclonal anti-(horseradish peroxidase) antibodies. Secondly, a fluorescein-labelled asialo- and digalactosylated biantennary complex sugar was synthesized and shown to bind to a Ricinus communis agglutinin column. This galactose-specific recognition was abolished by treating this fluorescein-labelled oligosaccharide with jack bean beta-galactosidase. Finally, two 1-N-glycyl beta-saccharide derivatives were modified with thiophosgene to form their corresponding isothiocyanate derivatives. Coupling of these isothiocyanate derivatives of sugars to BSA, amino-derivatized polystyrene plates and glass-fibre discs resulted in multiple sugar presentation. The binding of an anti-N-acetylglucosamine monoclonal antibody to N,N'-diacetylchitobiose residues presented on BSA and solid supports was shown by e.l.i.s.a. Similarly the binding of concanavalin A to asialo-, agalactosylated biantennary complex oligosaccharide residues attached to BSA was demonstrated by a competitive e.l.i.s.a. Our results demonstrate that N-linked neoglycoconjugates could be made readily available and they are valuable tools for the detailed analyses of carbohydrates and carbohydrate-binding proteins.
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PMID:Analysis of carbohydrate-protein interactions with synthetic N-linked neoglycoconjugate probes. 750 28

We studied the utilization of protein-hydrolyzed sweet cheese whey as a medium for the production of beta-galactosidase by the yeasts Kluyveromyces marxianus CBS 712 and CBS 6556. The conditions for growth were determined in shake cultures. The best growth occurred at pH 5.5 and 37 degrees C. Strain CBS 6556 grew in cheese whey in natura, while strain CBS 712 needed cheese whey supplemented with yeast extract. Each yeast was grown in a bioreactor under these conditions. The strains produced equivalent amounts of beta-galactosidase. To optimize the process, strain CBS 6556 was grown in concentrated cheese whey, resulting in a higher beta-galactosidase production. The beta-galactosidase produced by strain CBS 6556 produced maximum activity at 37 degrees C, and had low stability at room temperature (30 degrees C) as well as at a storage temperature of 4 degrees C. At -4 degrees C and -18 degrees C, the enzyme maintained its activity for over 9 weeks.
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PMID:Utilization of protein-hydrolyzed cheese whey for production of beta-galactosidase by Kluyveromyces marxianus. 1051 Apr 85

We previously clarified that the chitinase from the hyperthermophilic archaeon Thermococcus kodakaraensis KOD1 produces diacetylchitobiose (GlcNAc(2)) as an end product from chitin. Here we sought to identify enzymes in T. kodakaraensis that were involved in the further degradation of GlcNAc(2). Through a search of the T. kodakaraensis genome, one candidate gene identified as a putative beta-glycosyl hydrolase was found in the near vicinity of the chitinase gene. The primary structure of the candidate protein was homologous to the beta-galactosidases in family 35 of glycosyl hydrolases at the N-terminal region, whereas the central region was homologous to beta-galactosidases in family 42. The purified protein from recombinant Escherichia coli clearly showed an exo-beta-D-glucosaminidase (GlcNase) activity but not beta-galactosidase activity. This GlcNase (GlmA(Tk)), a homodimer of 90-kDa subunits, exhibited highest activity toward reduced chitobiose at pH 6.0 and 80 degrees C and specifically cleaved the nonreducing terminal glycosidic bond of chitooligosaccharides. The GlcNase activity was also detected in T. kodakaraensis cells, and the expression of GlmA(Tk) was induced by GlcNAc(2) and chitin, strongly suggesting that GlmA(Tk) is involved in chitin catabolism in T. kodakaraensis. These results suggest that T. kodakaraensis, unlike other organisms, possesses a novel chitinolytic pathway where GlcNAc(2) from chitin is first deacetylated and successively hydrolyzed to glucosamine. This is the first report that reveals the primary structure of GlcNase not only from an archaeon but also from any organism.
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PMID:Characterization of an exo-beta-D-glucosaminidase involved in a novel chitinolytic pathway from the hyperthermophilic archaeon Thermococcus kodakaraensis KOD1. 1292 90

An intracellular beta-glycoside hydrolase with beta-glucosidase and beta-galactosidase activity, designated beta-glucosidase BGL1, was isolated to apparent homogeneity from the thermophilic ascomycete Talaromyces thermophilus CBS 236.58. The monomeric enzyme has a molecular mass of 50 kDa (SDS-PAGE) and an isoelectric point of 4.5-4.6. The enzyme is active with both glucosides such as cellobiose and galactosides including lactose; based on the catalytic efficiencies determined glucosides are the preferred substrates. beta-Galactosidase activity of BGL1 is activated by various mono and divalent cations including Na+, K+ and Mg2+, and it is moderately inhibited by its reaction products glucose and galactose. Its pH optimum for the hydrolysis of galactosides is in the range of 5.5-6.0, and its optimum temperature was found to be 50 degrees C (15 min assay). In addition to its hydrolytic activity, BGL1 shows a significant transferase activity which results in the formation of galacto-oligosaccharides. These have recently attracted interest because of possible applications in food industry. The highest yields of oligosaccharides was approximately 20% when using 38 gl(-1) lactose as the starting material.
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PMID:Purification and characterisation of an intracellular enzyme with beta-glucosidase and beta-galactosidase activity from the thermophilic fungus Talaromyces thermophilus CBS 236.58. 1644 2

The yeast Kluyveromyces marxianus presents several interesting features that make this species a promising industrial yeast for the production of several compounds. In order to take full advantage of this yeast and its particular properties, proper tools for gene disruption and metabolic engineering are needed. The Cre-loxP system is a very versatile tool that allows for gene marker rescue, resulting in mutant strains free of exogenous selective markers, which is a very important aspect for industrial application. As the Cre-loxP system works in some non-conventional yeasts, namely Kluyveromyces lactis, we wished to know whether it also works in K. marxianus. Here, we report the validation of this system in K. marxianus CBS 6556, by disrupting two copies of the LAC4 gene, which encodes a beta-galactosidase activity.
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PMID:Application of the Cre-loxP system for multiple gene disruption in the yeast Kluyveromyces marxianus. 1762 62

Strains belonging to the yeast species Kluyveromyces marxianus have been isolated from a great variety of habitats, which results in a high metabolic diversity and a substantial degree of intraspecific polymorphism. As a consequence, several different biotechnological applications have been investigated with this yeast: production of enzymes (beta-galactosidase, beta-glucosidase, inulinase, and polygalacturonases, among others), of single-cell protein, of aroma compounds, and of ethanol (including high-temperature and simultaneous saccharification-fermentation processes); reduction of lactose content in food products; production of bioingredients from cheese-whey; bioremediation; as an anticholesterolemic agent; and as a host for heterologous protein production. Compared to its congener and model organism, Kluyveromyces lactis, the accumulated knowledge on K. marxianus is much smaller and spread over a number of different strains. Although there is no publicly available genome sequence for this species, 20% of the CBS 712 strain genome was randomly sequenced (Llorente et al. in FEBS Lett 487:71-75, 2000). In spite of these facts, K. marxianus can envisage a great biotechnological future because of some of its qualities, such as a broad substrate spectrum, thermotolerance, high growth rates, and less tendency to ferment when exposed to sugar excess, when compared to K. lactis. To increase our knowledge on the biology of this species and to enable the potential applications to be converted into industrial practice, a more systematic approach, including the careful choice of (a) reference strain(s) by the scientific community, would certainly be of great value.
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PMID:The yeast Kluyveromyces marxianus and its biotechnological potential. 1842 4

Klebsiella oxytoca (NRRL-B199), although able to produce 2, 3-butanediol from glucose, converted lactose mainly into acetic acid. By addition of a preparation of lactase (beta-galactosidase, EC 3.2.1.23), the fermentation of lactose in a stirred vessel was three-times faster and resulted in a high concentration of 2, 3-butanediol. The lactase confined in dead cells of Kluyveromyces lactis (CBS 683) was prepared by permeabilization with solvents and fixation with glutaraldehyde. The cells were coimmobilized by adhesion to glass wool after treatment of the latter with chitosan, which ensured cell-support electrostatic attraction. The cell loading (dry weight) was ca. 9 gL(-1) for the yeast and ca. 2 gL(-1) for the bacteria. In the presence of culture medium, the adhesion of both cells was stable and the bacteria tended to form biofilms. The stability of the coimmobilized cells was demonstrated by the continous conversion of lactose into 2, 3-butanediol at 30oC during 25 days. The coimmobilization system gave output concentrations (14 gL(-1)) and rate of production (1 gL(-1) h(-1)) of 2, 3-butanediol from lactose, similar to those obtained in the literature with immobilized cells and glucose. Compared to the literature data on direct conversion of lactose using pure cultures, the present results showed higher butanediol concentrations and 10 to 100 times higher rates of production.
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PMID:Co-immobilization by adhesion of beta-galactosidase in nonviable cells of Kluyveromyces lactis with Klebsiella oxytoca: conversion of lactose into 2, 3-butanediol. 1858 71

We used preS2-S'-beta-galactosidase, a three domain fusion protein that aggregates extensively at 43 degrees C in the cytoplasm of Escherichia coli to search for multicopy suppressors of protein aggregation and inclusion bodies formation, and took advantage of the known differential solubility of preS2-S'-beta-galactosidase at 37 and 43 degrees C to develop a selection procedure for the gene products that would prevent its aggregation in vivo at 43 degrees C. First, we demonstrate that the differential solubility of preS2-S'-beta-galactosidase results in a lactose-positive phenotype at 37 degrees C as opposed to a lactose-negative phenotype at 43 degrees C. We searched for multicopy suppressors of preS2-S'-beta-galactosidase aggregation at 43 degrees C by selecting pink lactose-positive colonies on a background of white lactose-negative colonies after transformation of bacteria with an E. coli gene bank. We found only two multicopy suppressors of preS2-S'-beta-galactosidase aggregation at 43 degrees C, protein isoaspartate methyltransferase (PIMT) and the membrane components ChbBC of the N,N'-diacetylchitobiose phosphotransferase transporter. We have previously shown that PIMT overexpression reduces the level of isoaspartate in preS2-S'-beta-galactosidase, increases its thermal stability and consequently helps in its solubilization at 43 degrees C (Kern et al., J. Bacteriol. 187, 1377-1383). In the present work, we show that ChbBC overexpression targets a fraction of preS2-S'-beta-galactosidase to the membrane, and decreases its amount in inclusion bodies, which results in its decreased thermodenaturation and in a lactose-positive phenotype at 43 degrees C. Cross-linking experiments show that the inner membrane protein ChbC interacts with preS2-S'-beta-galactosidase. Our results suggest that membrane docking of aggregation-prone proteins might be a useful method for their solubilization.
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PMID:Membrane docking of an aggregation-prone protein improves its solubilization. 1880 75

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