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)

We have isolated two phenotypically distinct nonfastidious Francisella strains (Fx1 and Fx2) from the blood of compromised patients with pneumonia and compared them with eight other Francisella strains, including Francisella tularensis biovar tularensis, F. tularensis biovar novicida, and F. philomiragia. Our isolates grew well on sheep blood agar, chocolate agar, modified Thayer-Martin agar, and Trypticase soy agar. Fx1 and Fx2 were determined to be within the Francisella genus by cellular fatty acid analysis and by the utilization of glucose, production of H2S and catalase, and lack of motility, oxidase, nitrate reductase, and gelatinase. They were additionally shown to belong to the species F. tularensis by sequencing of two variable regions comprising approximately 500 nucleotides of the 16S rRNA gene. Also, RNA probe hybridization confirmed their belonging to the species F. tularensis. However, the new strains, which are not identical, are distinguished from other F. tularensis strains by growth characteristics, repetitive extragenic palindromic PCR fragment pattern, and some biochemical tests. Key biochemical differences included the findings that Fx1 was positive for beta-galactosidase and arabinose hydrolysis and that both strains were citrulline ureidase positive and glycerol negative. Commercial F. tularensis antiserum agglutinated stock F. tularensis strains but not Fx1, Fx2, F. tularensis biovar novicida, or F. philomiragia; serum from either patient failed to agglutinate or only weakly agglutinated commercial antigen but showed agglutination when tested against each patient's respective isolate. Fx1 and Fx2 produced beta-lactamase. Because of their good growth, negative serology, and biochemical profile, the organisms could be misidentified in the clinical laboratory if standard strategies or commercial identification systems are used.
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PMID:Characterization of two unusual clinically significant Francisella strains. 881 97

We developed two Streptomyces-Escherichia coli shuttle vectors. The plasmid pRES102, consisting of the essential region of pRES1 and the thiostrepton resistance gene (tsr) fragment of pIJ702, was combined with the E. coli plasmid vector pUC18 or pUC19. The resulting shuttle vectors, designated pRES18 and pRES19, respectively, have relatively compact size (6.25 kb), low copy number, multiple cloning sites reciprocally arranged in opposite directions, and selection markers for both Streptomyces (tsr) and E. coli (beta-lactamase (bla) and beta-galactosidase (lacZ). These shuttle vectors are capable of carrying DNA fragments as long as 10 kb, of being maintained in S. griseus, S. lavendulae and S. lividans, and are compatible with pIJ702.
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PMID:Construction of pRES18 and pRES19, Streptomyces-Escherichia coli shuttle vectors carrying multiple cloning sites. 893 35

Here, we report on the construction of three integrative plasmids for Bacillus subtilis (Bs) allowing in vitro construction of transcriptional fusions. These plasmids contain a neomycin- or tetracycline-resistance cassette and one of three promoterless genes: bgaB (encoding beta-galactosidase), cat (chloramphenicol acetyltransferase), or xylE (catechol 2,3-dioxygenase). All cassettes are flanked by the 3'- and 5'-ends of the amyE gene (encoding alpha-amylase) allowing integration of these cassettes at the amyE locus of the Bs chromosome. For propagation and selection in Escherichia coli, the plasmids contain the pBR322 origin of DNA replication and the beta-lactamase-encoding bla gene. Four unique restriction sites can be used for insertion of restriction fragments carrying promoter fragments. All three reporter genes express heat-stable enzymes (stable up to at least 50 degrees C for 30 min) as shown here. We would like to point to the modular nature of these plasmids where the three reporter genes and the two resistance cassettes can be combined in any permutation. The versatility of the promoter-probe vectors was demonstrated by the integration of the promoters of the dnaK and groE operons of Bs and following their heat-inducible expression.
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PMID:Integrative vectors for constructing single-copy transcriptional fusions between Bacillus subtilis promoters and various reporter genes encoding heat-stable enzymes. 898 64

Here we report on the construction of two integrative plasmids for Bacillus subtilis allowing in vitro construction of translational fusions. Both plasmids contain two cassettes in tandem: the bgaB gene encoding a heat-stable beta-galactosidase devoid of its own regulatory sequences and the first two codons followed by a neomycin-resistance gene for selection in B. subtilis. Both cassettes are flanked by the 3'- and 5'-end of the amyE gene (encoding alpha-amylase) allowing integration of both cassettes at the amyE locus of the B. subtilis chromosome. For propagation in Escherichia coli, the plasmids contain the pBR322 origin of DNA replication and the beta-lactamase-encoding gene. Whereas one vector needs a promoter, a Shine-Dalgarno sequence and the beginning of a gene fused in-frame to bgaB, the other one already carries a constitutive promoter. The versatility of the gene fusion vectors was demonstrated by the integration of the regulatory regions of the dnaK and the cat-86 genes. In the first case, heat-inducible expression was found, and by comparison with an operon fusion, it seems that the dnaK operon is regulated at both the transcriptional and the posttranscriptional level. In the second case, chloramphenicol-inducible regulation of the gene fusion could be demonstrated.
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PMID:Integrative vector for constructing single-copy translational fusions between regulatory regions of Bacillus subtilis and the bgaB reporter gene encoding a heat-stable beta-galactosidase. 916 5

Homologues of the Na+/glucose cotransporter, the SGLT family, include sequences of mammalian, eubacterial, yeast, insect and nematode origin. The cotransported substrates are sugars, inositol, proline, pantothenate, iodide, urea and undetermined solutes. It is reasonable to expect that the SGLT family members share a similar or identical topology of membrane spanning elements, by virtue of their common ancestry and similar coupling of solute transport to downhill sodium flux. Here we examine their membrane topologies as deduced from diverse analyses of their primary sequences, and from their sequence correlations with the experimentally determined topology of the human Na+/ glucose contransporter SGLT1. Our analyses indicate that all family members share a common core of 13 transmembrane helices, but that some, like SGLT1 itself, have one additional span appended to the C-terminus, and still others, two. One bacterial member incorporates an additional span at the N-terminus. Sequence comparisons indicative of common ancestry of the SGLT and the [Na+ + Cl-] transporter families are introduced, and evaluated in light of their topologies. New evidence concerning the previously asserted common ancestry of SGLT1 and an N-acetylglucosamine permease of the bacterial phosphotransferase system is considered. Finally, we analyze observations which lead us to conjecture that the experimental strategy most commonly employed to reveal the topology of bacterial transporters (i.e., the fusion of reporter enzymes such as phoA alkaline phosphatase, beta-lactamase or beta-galactosidase, to progressively C-truncated fragments of the transporter) has often instead so perturbed local topology as to have entirely missed pairs of adjacent membrane spans.
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PMID:Membrane topology motifs in the SGLT cotransporter family. 930 6

Resistance to piperacillin in several isolates of Citrobacter freundii and Enterobacter cloacae was investigated and confirmed to occur at a frequency of 10(-7) to 10(-6). Development of resistance to piperacillin was significantly suppressed by tazobactam but not by clavulanic acid. To elucidate the mechanism by which resistance suppression occurs, the effect of piperacillin plus tazobactam on the induction of AmpC beta-lactamase was analyzed by monitoring the beta-galactosidase activity of an inducible ampC-lacZ gene fusion in Escherichia coli. The combination exerted no inhibitory effect on AmpC beta-lactamase induction. Tazobactam also had no effect on the accumulation of a key intermediate in the AmpC beta-lactamase induction pathway, 1,6-anhydromurotripeptide, in an ampD mutant strain of E. coli. However, the addition of tazobactam to liquid cultures of E. cloacae 40001 in the presence of piperacillin at four times the MIC caused a delay in the recovery of the culture to piperacillin-induced stress. At 16 times the MIC, a complete suppression of regrowth occurred. Analysis of culture viability on piperacillin plates showed that the culture recovery was due to growth by moderately resistant mutants preexisting in the cell population, which at 16 times the MIC became susceptible to the combination. Evidence from the kinetics of inhibition of the E. cloacae 40001 AmpC beta-lactamase by clavulanic acid, sulbactam, and tazobactam and from the effects of these drugs on the frequency of resistance to piperacillin suggests that the suppressive effect of tazobactam on the appearance of resistance is primarily mediated by the beta-lactamase inhibitory activity.
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PMID:Mechanism of suppression of piperacillin resistance in enterobacteria by tazobactam. 933 44

Isolates of Rhodococcus equi from pneumonic foals possess an 85- or 90-kb virulence-associated plasmid. A prominent, thermoregulated surface antigen, VapA, encoded by these plasmids is thought to be important in virulence. A 135-kb fragment containing the origin of replication of R. equi strain 103 virulence-associated plasmid (pOTS) was identified, sequenced, and its location identified. A simple R. equi-Escherichia coli shuttle plasmid (pRE-1) derived from the E. coli plasmid pACYC177 and the pOTS ori was developed. The plasmid transformed readily and was stable in either host and expressed kanamycin resistance but not beta-lactamase in R. equi. An improved 5.9-kb vector, pRE-7, was developed from pRE-1 and pBluescript. Subcloning of vapA into the multiple cloning site of the beta-galactosidase gene of pRE-7 resulted in weak expression of the gene both in E. coli and R. equi. The shuttle vector may be useful in examining regulation of virulence gene expression in R. equi.
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PMID:Development of a Rhodococcus equi-Escherichia coli plasmid shuttle vector. 943 20

Peptide nucleic acid (PNA) is a DNA mimic with attractive properties for developing improved gene-targeted antisense agents. To test this potential of PNA in bacteria, PNAs were designed to target the start codon regions of the Escherichia coli beta-galactosidase and beta-lactamase genes. Dose-dependent and specific gene inhibition was observed in vitro using low nanomolar PNA concentrations and in vivo using low micromolar concentrations. Inhibition was more efficient for a permeable E. coli strain relative to wild-type K-12. The potency of the anti-beta-lactamase PNAs was abolished by a six base substitution, and inhibition could be re-established using a PNA with compensating base changes. Antisense inhibition of the beta-lactamase gene was sufficient to sensitize resistant cells to the antibiotic ampicillin. The results demonstrate gene- and sequence-specific antisense inhibition in E. coli and open possibilities for antisense antibacterial drugs and gene function analyses in bacteria.
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PMID:Antisense inhibition of gene expression in bacteria by PNA targeted to mRNA. 955 26

Escherichia coli strain HB101 harboring an expression plasmid bearing calf prochymosin gene under the control of the tac promoter was grown in the presence of IPTG with or without novobiocin at 28 and 40 degrees C, respectively. The differential rates of synthesis of prochymosin inclusions, and, for comparison, of beta-lactamase and beta-galactosidase, as well as plasmid copy number, were determined during the first hours of steady state growth. At 28 degrees C the induced expression of prochymosin gene was almost blocked. Addition of novobiocin did not alleviate this effect. In fact, it strengthened it, and we conclude that both these additive inhibitory effects are a consequence of the decrease in negative superhelical tension of plasmid DNA to an insufficient level. At 40 degrees C the differential rate of prochymosin synthesis was markedly enhanced. Since the copy number of the expression plasmid increased approximately to the same extent, we conclude that an increase in gene dose is the cause. The stimulation of cloned heterologous gene expression at 40 degrees C and inhibition at 28 degrees C may be conveniently used in biotechnological-scale cultivations of some recombinant bacteria.
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PMID:Effects of temperature and novobiocin on the expression of calf prochymosin gene and on plasmid copy number in recombinant Escherichia coli. 956 30

A low-copy expression vector has been constructed from a 9 Kbp region of the Escherichia coli F plasmid containing the oriV and oriS origins of replication. This plasmid carries the beta-lactamase gene (Apr) and the araBAD promoter/araC regulator for arabinose-inducible gene expression. A derivative which carries a lacZ reporter gene was found to be stably maintained for at least 150 generations. A related multi-copy plasmid was stably maintained in arabinose-free medium, but no plasmid-bearing segregants remained after 60 generations when lacZ expression was induced. Induced expression resulted in 27% (multi-copy) and 12% (low-copy) decreases in growth rate. The uninduced levels of beta-galactosidase were 200 units (multi-copy) and 15 units (low-copy).
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PMID:Construction and characterization of F plasmid-based expression vectors. 1009 85


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