Document Details
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Document Type |
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Thesis |
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Document Title |
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Studying the effect of chromium on efficiency of L-asparaginase – producing bacteria دراسة تأثير الكروميوم على كفاءة البكتيريا في إنتاج انزيم الاسبراجينيز |
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Subject |
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Faculty of Sciences |
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Document Language |
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Arabic |
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Abstract |
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L-Asparaginase is a tetrameric protein, having anti leukemic activity on human cells. Reactive oxygen species (ROS) production is the central mechanism for Chromium genotoxicity, which was demonstrated through its ability to generate superoxide radical anions as a type of ROS. Therefore, screen and identify L- asparaginase bacteria and verify the effect of chromium CrO3- [Cr (VI)] on bacterial L-asparaginase production and activity were required. In this study, fifty isolates were obtained from the rhizospheres of four plants (Anethum graveolens, Portulaca oleracea, Eruca sativa and Phaseolus vulgaris), and one isolate was obtained from the soil in Algamom, Makkah, Saudi Arabia. The fifty bacterial isolates were screened for potential L-asparaginase production using a rapid assay plate method on M9 medium containing L-asparagine with phenol red as an indicator. Four isolates showed promising enzyme activity. Based on morphological, biochemical characteristics and 16S rRNA gene sequencing data. The most potent isolates were selected, identified and designated as Acinetobacter baumannii (HO6), Acinetobacter haemolyticus (FA3), Acinetobacter lwoffii (FA1) and Tatumella ptyseos (FA2). The sequences were submitted to the NCBI GenBank under accession numbers, KY313625 for A. lwoffii (FA1), KY313626 for T. ptyseos (FA2), KY313627 for A. haemolyticus (FA3) and KY313633 for A. baumannii (HO6) isolates. FA2 is the first T. ptyseos strain was identified with high levels of L-asparaginase enzyme production. Moreover, the effect of antibiotics on bacterial growth of each strains as a sort of identification was examined and results showed that all strains were sensitive to Streptomycen, Levofloxacin, Tigecycline, Tobramycin, Vancomycin and Clindemycin, whereas were resistance to Ampicillin/Sulbactam and Metronidazole. In addition, another identification was the plasmid profile where results showed that the three strains harbored one plasmid whereas A. lwoffii (FA1) harbored two plasmids. Furthermore, these strains were used to examine the toxic effects of low concentrations of chromium (VI) on enzyme activity. Spectrophotometric analysis revealed that the three CrO3 [Cr(VI)] concentrations (0.05, 0.1 and 0.15 mM) reduced the enzyme activity of A. baumannii (HO6) and T. ptyseos (FA2) whereas enhanced the enzyme activity of A. haemolyticus (FA3). Therefore, A. baumannii (HO6) was selected to examine the toxic effects of low concentrations of chromium (VI) on L-asparaginase gene expression using quantitative real time PCR (qRT PCR) to verify of L-asparaginase gene expression levels in A. baumannii (HO6) before and after treatment with three different CrO3 [Cr(VI)] concentrations. Quantitative analysis of asparaginase gene mRNA showed that addition of three CrO3 [Cr(VI)] concentrations (0.05, 0.1 and 0.15 mM) reduced significantly the expression level of L-asparaginase gene compared with control where (P-value < 0.05). Also, the plasmid content within A. baumannii (HO6) was examined before and after treatment with the three chromium concentrations and the results showed plasmid curing. |
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Supervisor |
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Dr. Alawiah Mohammad AL-Hebshi |
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Thesis Type |
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Master Thesis |
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Publishing Year |
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1439 AH
2018 AD |
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Added Date |
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Thursday, February 1, 2018 |
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Researchers
| فضا مهجع العنزي | AL-Enazi, Fada Mohajji | Researcher | Master | |
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