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Biodegradation of 17alfa-ethinylestradiol by enzymes of ligninolytic fungi
Přenosilová, Lenka ; Stiborová, Marie (advisor) ; Černá, Věra (referee)
This work is aimed at the study of the effect of 17α-ethinylestradiol (EE2) on the production and characteristics of ligninolytic enzymes (laccase, Mn-dependent peroxidase and lignin peroxidase) in I. lacteus, T. versicolor, P. chrysosporium and P. ostreatus cultures grown on two types of liquid media. Enzyme activity production in fungal cultures was affected by the composition of culture medium. In the case of P. chrysosporium, the addition of EE2 to the complex- medium cultures led to a MnP activity stimulation and simultaneously LiP production was partially repressed in these cultures. In the mineral MM medium, no effect of EE2 on enzyme production by P. chrysosporium was observed. In EE2 treated MM cultures of P. ostreatus lower MnP activities were found when compared to biotic controls. In the case of T. versicolor cultures, the addition of EE2 to the complex medium caused laccase and LiP stimulation in the cultures. In the MM medium, however, only laccase production was affected by EE2. I. lacteus MnP production was partially repressed by EE2 in MM medium. In contrast to that, significantly higher MnP activities were detected in complex- medium I. lacteus cultures after the treatment with EE2. Further EE2 degradation by the fungal cultures was studied. The highest degradation effeciency was...
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Laccase activity profiling in Trametes versicolor cultures degrading endocrine-disrupting compound Delor 103
Plačková, Martina ; Svobodová, Kateřina (advisor) ; Mikušová, Gabriela (referee)
In this work endocrine disrupting potential of Delor 103, a commercial mixture of PCB congeners, was studied along with its effect on production of laccase by the ligninolytic fungus Trametes versicolor. Using a gene-reporter yeast assay for evaluation of hormonal activity Delor 103 showed an androgenic activity with an EC50 value of 2.29. 10-2 mg/l. Chlorbenzoic acids, Delor 103 potential metabolites resulting from microbial degradation, displayed on the other hand an estrogenic activity, indicating possible changes in hormonal activity of Delor 103 during its microbial degradation. The addition of Delor 103 to mineral medium T. versicolor cultures resulted in an up to 257times higher laccase activities detected in fungal cultures. Delor 103 induced enzymes showed different pI values from those of control cultures. In a complex malt-extract glucose medium (MEG) the stimulation effect of Delor 103 was kept down. Further, the production of laccase and synthesis of different pI forms depended strongly on the growth phase of fungal cultures. Exponencially growing cultures of T. versicolor were able to produce up to 7 different pI forms of laccase in responce to Delor 103 whereas stationary cultures produced only 4 enzyme forms with higher pI values. Stimulation of laccase activities in T. versicolor,...
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The effect of cancerogenic azo dye Sudan I on expression of biotransformation enzymes
Hejduková, Žaneta ; Dračínská, Helena (referee) ; Svášková, Dagmar (advisor)
Sudan I is a widely used azo dye which has the ability to cause carcinomas in organs and tissues of experimental animals. During reactions catalyzed by microsomal monooxygenase enzyme systems or cytoplasmic biotransformation enzymes, Sudan I is oxidized to reactive metabolites that covalently bind to nucleic acids and cause their damage. Sudan I can also be metabolized by reduction, e. g. by a DT-diaphorase enzyme (NQO1). Reduction of Sudan I is considered to be a detoxification reaction. In this work, the in vivo action of Sudan I is examined in terms of its ability to induce an expression of the biotransformation enzyme DT-diaphorase in tissues of rats treated with the azo dye. The aim of this work was to quantify the degree of NQO1 induction at mRNA level. After the isolation of total RNA from organs of rats treated with Sudan I, the RNA was converted to cDNA by reverse transcription using random hexamers as primers. Using specific probes, the abundance of mRNA for the enzyme NQO1 in the organs of treated rats was quantified by "real-time" PCR, relatively to the control gene with a constant expression (β-actin). Through comparing thus determined amounts of mRNA in individual organs of treated and untreated rats, it has been found that Sudan I had caused a significant increase in the expression...
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Vliv polynenasycených mastných kyselin n-3 na expresi vybraného genu u modelového organizmu
Strouhalová, Eliška
The aim of my thesis was to assess the influence of polyunsaturated fatty acids n 3 (DHA and EPA) on gene expression, mainly PPARy, in model organism. The expression of PPARy gene is increased by these polyunsaturated fatty acids and thus they decrease the risk of development of inflammation and atherosclerosis. This hypothesis was tested on 40 test rats (Wistar albino). The first 7 weeks the rats were fed with a diet containing beef tallow and sweet condensed milk until they manifested mild obesity and inflammation. During other 7 weeks they were divided into 4 groups, each containing 10 individuals. One of the groups was further fed with diet containing beef tallow (control group), other groups received diet with 6% content of oil from safflower added, 6% content of fish oil (high content of EPA) or 6% content of oil from Schizochytrium alga (high content of DHA). The expression of PPARy was measured by using molecular-biology methods at the end of the trial. It was proven that the diet rich on DHA increases PPARy gene expression (P<0,05). The increase of PPARy gene expression in a diet containing fish oil was inconclusive (P>0,05). With these results we confirmed the hypothesis that diet rich on PUFA n3 decreases the risk of inflammation development and atherosclerosis and that DHA is significantly more effective.
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Microarray Data Interpretation
Ludwig, Petr ; Šilhavá, Jana (referee) ; Smrž, Pavel (advisor)
This Bachelor thesis explains the basics of biochip or microarray data interpretation, starting with short introduction to genetics, especially genetic information significance evaluating. The focus was set mainly on the set of scripts transforming and analyzing the sample data. The data used in this thesis are a result of biochip analysis of the Colon Tumor tissues. The secondary result represents disclosing the main marker for this particular type of cancer, the primary result is evaluation of marker significance in the context of signaling pathways. The resulting pathways are sorted by relevance.
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Functional analysis of HSP complex expression in response to cold in \kur{Drosophila melanogaster}
ŠTĚTINA, Tomáš
The constitutively obsereved up-regulation of Hsp70 expression often led to premature conclusions about its critical role as a repair mechanism of cold injury that is, besides, expressed by protein misfolding/denaturation. In this study, we analyze the cold tolerance and the expression of 24 different mRNA transcripts of Hsp complex and other genes, that are associated with the repair of injury caused by cold. We use two strains of D. melanogaster: the wild type and the mutant type Hsp70- null, that lacks all 6 copies of the gene hsp70. We found out, that the larvae of two strains do not differ in their patterns of target genes expression during long term acclimation nor during recovery from chronic cold exposure and acute cold shock, therefore there is no transcriptional compensation of any other Hsp gene for the missing hsp70 in Hsp70- strain. The cold tolerance of Hsp70- strain larvae was impaired only, when they were exposed to strong acute cold shock by temperatures below -8°C. No difference in cold tolerance was observed, when the larvae were exposed to chronic cold exposure in 0°C or to mild acute cold shock by temperatures up to -4°C. Based on our results we assess, that the cold injury caused by strong acute cold schock is of another nature than caused by mild cold conditions and only in the first case Hsp70 expression is critical for the repair of cold injury in Drosophila melanogaster larvae.
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