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)

The ilvBNC operon of Corynebacterium glutamicum encodes acetohydroxy acid synthase and isomero-reductase, which are key enzymes of L-isoleucine, L-valine and L-leucine syntheses. In this study we identified the transcript initiation site of ilvBNC operon 292 nucleotides in front of the first structural gene, and detected the formation of a short transcript from the leader region in addition to the full-length transcript of the operon. This identifies the control of ilvBNC transcription by an attenuation mechanism involving antitermination. Mutations in the leader region were made and their effect on the operon expression in ilvB'lacZ fusions was quantified. Although a presumed leader-peptide-coding region is only one nucleotide away from the transcript initiation site determined, there is clear evidence to support the formation of this leader peptide: (i) the substitution of initiation codon ATG of the peptide by AGG reduced lacZ expression of the appropriate fusion construct to 19%; (ii) the replacement of three subsequent Val codons by Ala codons resulted in the loss of Val-dependent expression; and (iii) a leader peptide LacZ fusion resulted in active beta-galactosidase. Based on these results, it is concluded that transcription of ilvBNC is controlled by a translational-coupled attenuation mechanism. The absence of a ribosome binding site for leader peptide formation means that additional mechanisms may contribute to the transcription control at the decoding initiation step in the leader peptide formation.
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PMID:Attenuation control of ilvBNC in Corynebacterium glutamicum: evidence of leader peptide formation without the presence of a ribosome binding site. 1623 99

It is shown here that Escherichia coli beta-galactosidase has a second Mg2+ binding site that is important for activity. Binding of Mg2+ to the second site caused the k(cat) (with oNPG as the substrate) to increase about 100 s(-1); the Km was not affected. The Kd for binding the second Mg2+ is about 10(-4)M. Since the concentration of free Mg2+ in E. coli is about 1-2 mM, the second site is physiologically significant. Non-polar substitutions (Ala or Leu) for Glu-797, a residue in an active site loop, eliminated the k(cat) increase. This indicates that the second Mg2+ site is near to Glu-797. The Ki values of transition state analogs were decreased by small but statistically significant amounts when the second Mg2+ site was occupied and Arrhenius plots showed that less entropic activation energy is required when the second site is occupied. These inhibitor and temperature results suggest that binding of the second Mg2+ helps to order the active site for stabilization of the transition state.
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PMID:Beta-galactosidase (Escherichia coli) has a second catalytically important Mg2+ site. 1712 92

High Mobility Group A (HMGA) is a family of architectural nuclear factors which play an important role in neoplastic transformation. HMGA proteins are multifunctional factors that associate both with DNA and nuclear proteins that have been involved in several nuclear processes including transcription. HMGA localization is exclusively nuclear but, to date, the mechanism of nuclear import for these proteins remains unknown. Here, we report the identification and characterization of a nuclear localization signal (NLS) for HMGA2, a member of the HMGA family. The NLS overlaps with the second of the three AT-hooks, the DNA-binding domains characteristic for this group of proteins. The functionality of this NLS was demonstrated by its ability to target a heterologous beta-galactosidase/green fluorescent protein fusion protein to the nucleus. Mutations to alanine of basic residues within the second AT-hook resulted in inhibition of HMGA2 nuclear localization and impairment of its function in activating the cyclin A promoter. In addition, HMGA2 was shown to directly interact with the nuclear import receptor importin-alpha2 via the second AT-hook. HMGA proteins are overexpressed and rearranged in a variety of tumors; our findings can thus help elucidating their role in neoplastic transformation.
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PMID:The second AT-hook of the architectural transcription factor HMGA2 is determinant for nuclear localization and function. 1732 44

Bacillus subtilis has various cell wall hydrolases, however, the functions and hydrolase activities of some enzymes are still unknown. B. subtilis CwlK (YcdD) exhibits high sequence similarity with the peptidoglycan hydrolytic L,D-endopeptidase (PLY500) of Listeria monocytogenes phage and CwlK has the VanY motif which is a D-alanyl-D-alanine carboxypeptidase (Pfam: http://www.sanger.ac.uk/Software/Pfam/). The beta-galactosidase activity observed on cwlK-lacZ fusion indicated that the cwlK gene was expressed during the vegetative growth phase, and Western blotting suggested that CwlK seems to be localized in the membrane. Truncated CwlK fused with a histidine-tag (h-DeltaCwlK) was produced in Escherichia coli and purified on a nickel column. The h-DeltaCwlK protein hydrolyzed the peptidoglycan of B. subtilis, and the optimal pH, temperature and NaCl concentration for h-DeltaCwlK were pH 6.5, 37 degrees C, and 0 M, respectively. Interestingly, h-DeltaCwlK could hydrolyze the linkage of L-alanine-D-glutamic acid in the stem of the peptidoglycan, however, this enzyme could not hydrolyze the linkage of D-alanine-D-alanine, suggesting that CwlK is an L,D-endopeptidase not a D,D-carboxypeptidase. CwlK could not hydrolyze polyglutamate from B. natto or peptidoglycan of Staphylococcus aureus. This is the first report describing the characterization of an L,D-endopeptidase in B. subtilis and also the first report in bacteria of the characterization of a PLY500 family protein encoded in chromosomal DNA.
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PMID:Characterization of new L,D-endopeptidase gene product CwlK (previous YcdD) that hydrolyzes peptidoglycan in Bacillus subtilis. 1758 76

Previous work has shown that the mechanosensitive (MS) channel of large conductance (MscL) is essential for preventing lysis of Bacillus subtilis log phase cells upon a rapid, severe osmotic downshock. Growing cells of B. subtilis strains lacking MscL and one or more putative MS channel proteins of small conductance (YhdY, YkuT and YfkC) showed even higher sensitivity to an osmotic downshock. The effect was greatest for a strain lacking MscL and YkuT, and a strain lacking all four MS channel proteins had a similar phenotype. These defects were complemented by expression of either MscL or YkuT in trans. All MS channel mutant strains ultimately became resistant to osmotic downshock in stationary phase but at varying times, with mscL ykuT strains taking the longest time to become resistant. Expression of beta-galactosidase from gene fusions to lacZ showed modest expression of ykuT and lower levels of expression of yhdY and yfkC when strains were grown in medium containing high salt. Sporulation of all MS channel mutant strains was normal, and the mutant spores germinated normally with L-alanine or dodecylamine.
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PMID:Growth, osmotic downshock resistance and differentiation of Bacillus subtilis strains lacking mechanosensitive channels. 1766 70

The inability to infect insect cell cultures at the highest achievable cell densities has imposed major limitations to both the fundamental understanding of the Baculovirus Expression Vector System (BEVS) as well as full exploitation of its potential productive capacity for recombinant (beta-galAcNPV) products. The current literature does not characterize and identify the exact nature of the observed limitations, which therefore has become the major objective and contribution of the following study. Critical densities for infection of Spodoptera frugiperda (Sf9) cells with nuclear polyhedrosis virus expressing beta-galactosidase (Autographa californica) grown in media both containing fetal calf serum (FCS) and free of serum were found to be at 2 x 10(6) and 5 x 10(6) cells/ml respectively. Medium exchange was found to completely reverse the effect if renewed up to 24 hours post-infection (HPI). The inevitable arrest of uninfected cell growth and decreased production of recombinant products at high cell densities of infection were both correlated to nutrient depletion. Cystine was found to be depleted in uninfected insect cell cultures at the onset of the stationary phase and in serum-free insect cell cultures infected with baculovirus above a cell density of 5 x 10(6) cells/ml. Neither glucose depletion nor accumulation of possible inhibitory metabolites such as alanine, ammonia, or lactate could be correlated to growth arrest or decreased recombinant product yields.
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PMID:Substrate limitation in the baculovirus expression vector system. 1863 7

The therapeutic usefulness of the quinoxaline derivatives XK469 (2-{4-[(7-chloro-2-quinoxalinyl)oxy]phenoxy}propionic acid) and SH80 (2-{4-[(7-bromo-2-quinolinyl)oxy]phenoxy}propionic acid) has been attributed to their abilities to induce G(2)/M arrest and apoptotic or autophagic cell death. Concentrations of XK469 or SH80 > or = 5 microM were cytostatic to cultures of the normal murine melanocyte cell line Melan-a. Higher concentrations caused dose-dependent cytotoxicity. Concentrations > or =10 microM provoked dramatic morphological changes typified by marked increases in cell size and granularity. XK469/SH80-treated cultures accumulated tetraploid (4N) DNA-containing cells within 24 h of treatment, an 8N population within 3 days, and a 16N population within 5 days. Increases in ploidy correlated with the appearance of multinucleated cells. Under no circumstances did cells exhibit evidence of furrow formation. Both drugs suppressed cytokinesis in additional mammalian cell lines. Cytotoxic concentrations of XK469 elevated DEVDase activities (a measure of procaspase-3/7 activation) and enhanced cellular staining by a fluorescent analog of the pan caspase inhibitor valine-alanine-aspartic acid-fluoromethyl ketone within 48 to 96 h of treatment. Within 48 h of treatment, cytostatic and cytotoxic concentrations of XK469 elevated p21 contents, reduced Bcl-2 and Bcl-XL contents, and induced autophagy, as monitored by the accumulation of phosphatidylethanolamine-modified cleavage product of microtubule-associated protein light chain 3 (LC3-II). Cultures treated with > or =10 microM XK469 or SH80 for 5 days could not be induced to divide upon removal of drugs. Such cultures maintained high LC3-II contents, exhibited reduced cyclin E and D1 contents, and extensively expressed senescence-associated beta-galactosidase within 14 to 17 days of cessation of drug treatment. Hence, XK469 and SH80 inhibit cytokinesis, promote polyploidy, and induce senescence in Melan-a cells.
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PMID:The chemotherapeutic agents XK469 (2-{4-[(7-chloro-2-quinoxalinyl)oxy]phenoxy}propionic acid) and SH80 (2-{4-[(7-bromo-2-quinolinyl)oxy]phenoxy}propionic acid) inhibit cytokinesis and promote polyploidy and induce senescence. 1906 41

A Gram-positive, short diphtheroid-shaped organism was isolated from a sow's placenta of an abortion. This novel isolate, strain Murakami(T), was examined physiologically, chemotaxonomically and phylogenetically. Cells had an irregular V-shaped or palisade arrangement. Colonies appeared translucent on TMVL agar. Cells were strictly anaerobic, negative for catalase and gelatin decomposition and positive for nitrate reduction and soluble starch hydrolysis. Fourteen sugars including glucose were utilized as carbon sources for growth, but 15 sugars including arabinose were not. alpha-Galactosidase, beta-galactosidase, alpha-glucosidase and leucine arylamidase were produced, but beta-glucosidase was not. Fermentation products were lactic, succinic and acetic acids. Sugars of whole cells consisted of rhamnose and ribose. The amino-acid composition of the peptidoglycan was glutamic acid, alanine and lysine in the molar ratio of 1 : 2 : 1. The main fatty acid components of whole cells were C(14 : 0), C(16 : 0), C(16 : 1)omega7 and C(18 : 1)omega9. The bacterial menaquinone was MK-10(H(4)). The polar lipids were phosphatidylethanolamine and two unknown phosphatidylinositol mannosides. The G+C content of the genomic DNA of strain Murakami(T) was 63.8 mol%. Phylogenetic analysis of 16S rRNA gene sequences from strain Murakami(T) and other members of the genus Arcanobacterium supported the phenotypic findings that strain Murakami(T) represents a novel species, for which the name Arcanobacterium abortisuis sp. nov. is proposed. The type strain is Murakami(T) (=ATCC BAA-1522(T) =DSM 19515(T) =JCM 14813(T)).
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PMID:Arcanobacterium abortisuis sp. nov., isolated from a placenta of a sow following an abortion. 1950 37

Gram-positive, non-spore-forming rods were isolated from a human osteo-articular sample (strain 7400942(T)). Based on cellular morphology and the results of biochemical analysis, this strain was tentatively identified as a novel species of the genus Actinomyces. Phylogenetic analysis based on 16S rRNA gene sequence comparisons showed that the bacterium was closely related to the type strain of Actinomyces denticolens (96.9 % 16S rRNA gene sequence similarity). A comparison of biochemical traits showed that strain 7400942(T) was distinct from A. denticolens in a number of characteristics, i.e. in contrast with A. denticolens, strain 7400942(T) was negative for nitrate reduction and for beta-galactosidase, alpha-glucosidase and alanine arylamidase activities, it was positive for acid production from N-acetylglucosamine, melezitose and glycogen, and it was negative for acid production from turanose. Matrix-assisted laser-desorption/ionization time-of-flight MS protein analysis confirmed that strain 7400942(T) represents a novel species, as scores obtained for its spectra were significant (>2.2) only with strain 7400942(T). On the basis of phenotypic data and phylogenetic inference, it is proposed that this strain should be designated Actinomyces timonensis sp. nov.; the type strain is strain 7400942(T) (=CSUR P35(T)=CCUG 55928(T)).
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PMID:Actinomyces timonensis sp. nov., isolated from a human clinical osteo-articular sample. 1968 13

It has long been believed that amino acids comprising proteins of all living organisms are only of the L-configuration, except for Gly. However, peptidyl D-amino acids were observed in hydrolysates of soluble high molecular weight fractions extracted from cells or tissues of various organisms. This strongly suggests that significant amounts of D-amino acids are naturally present in usual proteins. Thus we analyzed the D-amino acid contents of His-tag-purified beta-galactosidase and human urocortin, which were synthesized by Escherichia coli grown in controlled synthetic media. After acidic hydrolysis for various times at 110 degrees C, samples were derivatized with 4-fluoro-7-nitro-2, 1, 3-benzoxadiazole (NBD-F) and separated on a reverse-phase column followed by a chiral column into D- and L-enantiomers. The contents of D-enantiomers of Ala, Leu, Phe, Val, Asp, and Glu were determined by plotting index D/(D + L) against the incubation time for hydrolysis and extrapolating the linear regression line to 0 h to eliminate the effect of racemization of amino acids during the incubation. Significant contents of D-amino acids were reproducibly detected, the D-amino acid profile being specific to an individual protein. This finding indicated the likelihood that D-amino acids are in fact present in the purified proteins. On the other hand, the D-amino acid contents of proteins were hardly influenced by the addition of D- or L-amino acids to the cultivation medium, whereas intracellular free D-amino acids sensitively varied according to the extracellular conditions. The origin of these D-amino acids detected in proteins was discussed.
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PMID:Detection of D-amino acids in purified proteins synthesized in Escherichia coli. 1976 21


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