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)

To examine the role of reactive oxygen species on the invasive phenotype of cancer cells, we overexpressed manganese- and copper-zinc-containing superoxide dismutases (MnSOD, CuZnSOD) and catalase (Cat) in hamster cheek pouch carcinoma (HCPC-1) cells in vitro using adenoviral vector-mediated gene transfer. Hamster cheek pouch carcinoma cells were transduced with these adenoviral vector constructs alone, or in combination, at concentrations [i.e., multiplicity of infectivity (MOI)] of 100 MOI each. The Escherichia coli beta-galactosidase reporter construct was used as a control virus. Protein expression was examined by Western blot analysis and enzymatic activities were measured using spectrophotometry. To observe the effects of transgene overexpression on in vitro tumor cell invasion, we used the membrane invasion culture system, an accurate and reliable method for examining tumor cell invasion, in vitro. This assay measures the ability of tumor cells to invade a basement membrane matrix consisting of type IV collagen, laminin, and gelatin. MnSOD overexpression resulted in a 50% increase in HCPC-1 cell invasiveness (p < .001); co-overexpression of MnSOD with Cat partially inhibited this effect (p < .05). Moreover, co-overexpression of both SODs resulted in a significant increase in invasiveness compared with the parental HCPC-1 cells (p < .05). These changes could not be correlated with the 72 kDa collagenase IV or stromolysin activities using zymography, or the downregulation of the adhesion molecules E-cadherin or the alpha4 subunit of the alpha4beta1 integrin. These results suggest that hydrogen peroxide may play a role in the process of tumor cell invasion, but that the process does not rely on changes in matrix metalloproteinase activity in the cells, or the expression of cell adhesion molecules.
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PMID:Effects of antioxidant enzyme overexpression on the invasive phenotype of hamster cheek pouch carcinoma cells. 1049 Feb 77

A toxicity test using a transgenic strain of the free-living soil nematode Caenorhabditis elegans carrying a stress-inducible beta-galactosidase reporter has been adapted for use in soil biomonitoring. High concentrations (250 microg. g(-1)) of cadmium are required to induce the stress response in worms exposed to Lufa 2.2 soil. Even at relatively high concentrations, the response to copper and zinc additions alone is minimal, yet combinations of cadmium and copper in the test soil induce a larger response than with cadmium alone at the same concentration. In contrast, the addition of both zinc and cadmium induces a lower response than cadmium additions alone. Analysis of the interstitial water suggests that there is preferential occupation by copper of sorption sites in the soil, allowing more cadmium to remain in solution. Conversely, cadmium and zinc would appear to interact similarly with the soil constituents, resulting in an increase of both metals in solution with increased additions to the soil. Aquatic tests mimic the results of the soil test, so it is not increased cadmium availability alone that causes an increased stress response when both cadmium and copper are present. The presence of other metals could reduce the amount of cadmium available, which may be one factor in the zinc moderation of the stress response to cadmium. Intracellular mechanisms may also contribute to the copper enhancement of the stress response to cadmium.http://link. springer-ny.com/link/service/journals/00244/bibs/37n4p503.++ +html</HEA
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PMID:Effect of single and paired metal inputs in soil on a stress-inducible transgenic nematode. 1050 98

Angiotensin II stimulates vascular NADPH oxidase to produce superoxide, which can react with nitric oxide and impair vasomotor function. We tested the hypothesis that the overexpression of endothelial nitric oxide synthase (eNOS) or superoxide dismutase (SOD) would correct angiotensin II-induced endothelial dysfunction. We examined the effects of the gene transfer of eNOS or 2 isoforms of SOD to the aorta in angiotensin II-treated rabbits on vasomotor function. New Zealand White rabbits were treated for 1 week with angiotensin II (100 ng. kg(-1). min(-1)) or saline by osmotic minipumps. In angiotensin II-treated rabbits, mean blood pressure was 107+/-8 mm Hg; it was 67+/-5 mm Hg in saline-infused rabbits (P<0.05). In aortas from angiotensin II-treated rabbits, lucigenin-enhanced chemiluminescence demonstrated a 2.5-fold increase in superoxide levels, and the oxidative fluorescent probe hydroethidine indicated increased superoxide levels throughout the vascular wall, especially in the endothelium and adventitia. Maximal relaxation to acetylcholine was less in aortas from rabbits treated with angiotensin II (72+/-5% versus 87+/-4% in saline-treated rabbits; P<0.01), but responses to sodium nitroprusside were similar. Segments of the thoracic aorta were incubated in vitro with an adenoviral vector that expressed eNOS, copper zinc SOD (CuZnSOD), extracellular SOD (ECSOD), or beta-galactosidase. beta-Gal treatment with adenovirus containing the gene for eNOS (AdeNOS) but not adenovirus containing the gene for beta-gal (Adbeta-gal) (control virus) restored responses to acetylcholine (82+/-3% after AdeNOS and 67+/-4% after Adbeta-gal). Gene transfer of CuZnSOD or ECSOD did not improve the endothelium-dependent relaxation of the aorta in rabbits that received angiotensin II. Thus, gene transfer of eNOS, but not SOD, effectively restores vasomotor function in angiotensin II-infused rabbits.
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PMID:Gene transfer of endothelial nitric oxide synthase reduces angiotensin II-induced endothelial dysfunction. 1067 3

The functionality of beta-galactosidase encoded by the E. coli lacZ gene as a reporter of gene expression in C. glabrata was investigated. C. glabrata/E. coli shuttle vectors were constructed, containing both a C. glabrata CEN-ARS cassette, to allow regular segregation and episomal replication of the plasmids, and the lacZ coding sequence of E. coli. The functionality of beta-galactosidase in C. glabrata was verified by inserting the promoter and the 5' coding region of the HIS3 gene from C. glabrata directionally upstream of the lacZ gene. By fusing the promoter of the copper-controlled MTII gene to the lacZ reporter, we showed that beta-galactosidase activity can be differentially induced in C. glabrata. beta-galactosidase reporter activities were detected qualitatively by an indirect filter assay and quantitatively from permeabilized cells.
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PMID:Candida glabrata shuttle vectors suitable for translational fusions to lacZ and use of beta-galactosidase as a reporter of gene expression. 1076 36

The development of an expanded bed process for the direct extraction and partial purification of beta-galactosidase from unclarified Escherichia coli homogenates using its natural affinity for metal loaded STREAMLINE Chelating is described. Small packed beds were used to determine the effect of chelated metal ion (Cu2+, Ni2+, Co2+ or Zn2+), loading pH and ionic strength on the selective binding capacity, and recovery of beta-galactosidase from clarified homogenates. An elution protocol was developed using the competitive displacer, imidazole, to recover beta-galactosidase in 87% yield and 3.4-fold purification. These results were then used to develop a separation for the recovery of beta-galactosidase from unclarified homogenates in a 2.5-cm diameter expanded bed. Although Ni2+ loaded STREAMLINE Chelating had a 5% dynamic capacity for beta-galactosidase of just 118 U ml(-1) (0.39 mg ml(-1)), the low capacity was thought to be due to the large size of the target (464,000) relative to the exclusion limit of the macroporous adsorbent. Despite this low capacity, Ni2 STREAMLINE Chelating was used successfully to recover beta-galactosidase from an unclarified homogenate in 86.4% yield and at 5.95-fold purification. The degree of purification relative to a commercial standard, as assessed using the purification factor and sodium dodecyl sulphate-polyacrylamide gel electrophoresis was high suggesting that this pseudo-affinity procedure compared favourably with alternative methods.
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PMID:Immobilised metal affinity chromatography of beta-galactosidase from unclarified Escherichia coli homogenates using expanded bed adsorption. 1076 97

Three biochemical parameters, DNA quantification in soil samples and two enzymatic activities, beta-galactosidase and dehydrogenase have been assessed as potential end-points for the use in cost-effective toxicity tests on soil microorganisms. The assessment included the development of a classical dose-response 24-h assay and the incorporation of measurements of the effects on microbial activities in soil column leaching studies and multispecies miniaturised terrestrial systems (MTS). Four different chemicals, copper, a new herbicide, thiabendazole and fenthion were studied. A rapid fluorescence DNA quantification technique did not produce adequate responses. The efforts to quantify DNA after extraction and clean-up procedures failed due to the presence of humic acids. From the protocol of the technique one could see that the technical procedure is time-consuming and expensive and, for this reason, not suitable for use as a parameter in rapid and cost-effective tests. However, the enzymatic activities showed their potential as toxicity end-points. Copper produced a concentration/response inhibition of beta-galactosidase and dehydrogenase with EC50 values of 78.39 and 24.77 mg Cu/kg soil, respectively. In the soil column study, these endpoints allowed the measurement of the microbial activities through the column. The effects of the new herbicide on beta-galactosidase and dehydrogenase activities were statistically significant for the highest application dose (40 g/ha). Thiabendazole affected the microbial activity when mixed within the soil, but no effects were observed when this fungicide was applied on the soil surface. Fenthion produced effects when applied either in the soil or on the soil surface. These results can be explained by the low mobility of thiabendazole. The results show the capabilities of these biochemical parameters to be included as endpoints in cost-effective bioassays.
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PMID:Rapid and cost-effective multiparameter toxicity tests for soil microorganisms. 1080 43

Copper-induced metallothionein (MT) synthesis in Saccharomyces cerevisiae was investigated in order to associate this exclusively with Cu2+ in vivo, when cultured in nutrient medium containing other heavy metal ions. Expression of the CUP1 promoter/lacZ fusion gene was inhibited by all heavy metal ions tested, especially Cd2+ and Mn2+. By adding Cd2+ and Mn2+ at 10 microM concentration, the beta-galactosidase activity decreased by about 80% and 50% of the maximum induction observed with 1 mM CuSO4, respectively. Furthermore, cell growth was markedly inhibited by combinations of 1 mM-Cu2+ and 1 microM-Cd2+. Therefore, the yeast S. cerevisiae could not rely on MT synthesis as one of the copper-resistance mechanisms, when grown in a Cd2+ environment. In contrast, the presence of Mn2+ in the nutrient medium showed alleviation rather than growth inhibition by high concentrations of Cu2+. The recovery from growth inhibition by Mn2+ was due to decreased Cu2+ accumulation. Inhibitory concentrations of Co2+, Ni2+ and Zn2+ on expression of the CUP1p/lacZ fusion gene were at least one order of magnitude higher than that of Cd2+ and Mn2+. These results are discussed in relation to Cu2+ transport and Cu-induced MT synthesis in the copper-resistance mechanism of the yeast S. cerevisiae.
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PMID:Effect of some heavy metal ions on copper-induced metallothionein synthesis in the yeast Saccharomyces cerevisiae. 1081 30

One way of improving heterologous protein production is to use high cell density systems, one of the most attractive being the flocculating yeast production system. Also, lactose is available in large amounts as a waste product from cheese production processes. The construction of flocculent and non-flocculent brewer's yeast strains secreting beta-galactosidase and growing on lactose is presented. A plasmid was constructed coding for an extracellular beta-galactosidase of Aspergillus niger and having, as selective marker, the yeast CUP1 gene conferring resistance to copper. This selective marker allows for the transformation of wild-type yeasts. This work represents an important step towards the study of heterologous protein secretion by flocculent cells.
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PMID:Construction of a flocculent brewer's yeast strain secreting Aspergillus niger beta-galactosidase. 1095 11

Downstream from the ptsHI operon of Lactobacillus sakei, the genes atkY and atkB, organized in an operon, were observed. The two putative proteins, AtkB and AtkY, show sequence similarity to the Enterococcus hirae copper P-type ATPase, responsible for copper efflux, and its negative regulator. Characterization of AtkB as a copper P-type ATPase could not be demonstrated since an atkB mutant did not show any phenotype. Thus, another strategy was followed in order to investigate the transcriptional regulation of the atkYB locus, leading to the development of new genetic tools for L. sakei. A plasmid was constructed, the use of which allowed gene replacement at the lacLM locus in L. sakei by two successive crossovers. A strain deleted of the lacLM operon encoding the beta-galactosidase of L. sakei was constructed by this method, and the Escherichia coli lacZ gene could then be used as a reporter gene to investigate the regulation of atkYB. Results show that the atkYB operon is induced by small concentrations of CuSO(4) (30 to 40 microM) but not when CuSO(4) is omitted or added at higher concentrations.
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PMID:Development of genetic tools for Lactobacillus sakei: disruption of the beta-galactosidase gene and use of lacZ as a reporter gene To study regulation of the putative copper ATPase, AtkB. 1101 Aug 70

Transcription of the ybcZ--ylcA ylcBCD--ybdE region of the Escherichia coli K38 chromosome was analysed by Northern RNA--DNA hybridization, RT-PCR and primer extension. Transcription of a dicistronic ybcZ--ylcA mRNA and a tetracistronic ylcBCD--ybdE mRNA was induced by silver and was initiated from the sigma-70 promoters ylcAp and ylcBp. Expression of beta-galactosidase activity from a Phi(ylcBp--lacZ) operon fusion was also induced by Ag(+) and Cu(2+), but not by Zn(2+). In-frame deletion of ybdE from the chromosome yielded a silver-sensitive E. coli mutant strain which did not differ in its copper resistance from its wild-type strain. On the other hand, deletion of the copA gene for the copper-exporting P-type ATPase CopA resulted in copper sensitivity, but not in silver sensitivity. A Delta ybdE Delta copA double mutant strain behaved towards copper as the Delta copA strain and towards silver as the Delta ybdE strain. Thus, in E. coli, the YlcBCD--YbdE system may be involved in silver- but not in copper resistance, and CopA may be involved in copper- but not in silver resistance.
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PMID:The product of the ybdE gene of the Escherichia coli chromosome is involved in detoxification of silver ions. 1128 92


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