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Keywords: pseudopartitives, agreement attraction, agreement
Published in RUNG: 22.09.2021; Views: 1176; Downloads: 47
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Abstract: Aims:Preparation of the new metals - polymeric composite, Met x-EPS (I), to be used as a green catalyst in water or in two-phase aqueous conditions. Study Design: Recovery and valorization of polymetallic wastes to obtain directly new catalysts using a microorganism to explore their application in removal of difficult and dangerous chemical pollutants present in aqueous environment
Keywords: Metals - polymeric composite, biogenerated polymetallic exopolysaccharide, new catalyst from metallic wastes, hydrodechlorination of PCBs in water
Published in RUNG: 06.05.2019; Views: 2906; Downloads: 0
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Abstract: The experimental work of this thesis was performed at the International Centre for Genetic Engineering and Biotechnology (ICGEB) in the Human Molecular Genetics Group, under the scientific direction of Prof. Franco Pagani. The project was developed during the academic years 2014-2017. Modified U1 RNAs, also named Exon Specific U1s (ExSpeU1s) represent a novel class of small RNA-based molecules that correct exons splicing defects. To evaluate their therapeutic potential focused on Familial Dysautonomia (FD), a rare autosomal recessive disorder characterized by progressive degeneration of the sensory and autonomic nervous system. More than 99% of patients are homozygous for the T to C transition in position 6 of the IKBKAP intron 20 (c.2204+6T>C). This substitution modifies the exon 20 5’ splice site (5’ss) inducing exon skipping in a tissue-specific manner and reducing the total amount of IKAP protein. The molecular mechanisms underlying the IKBKAP mis-splicing are not completely clear and there are no effective treatments. In this thesis, I investigated the therapeutic potential of ExSpeU1s and the role of cis- and trans-acting factors that regulates IKBKAP splicing. Using a splicing functional assay, I identified ExSpeU1s that bind to intron 20 sequences and rescue the exon 20 skipping defect. Interestingly, their rescue activity was modulated by several splicing factors and requires a critical exonic splicing enhancer element. Transfection experiment showed the involvement of both enhancing (TIA1, PTBP1 and PTB4) and inhibitory (SRSF3, hnRNPA1, FOX and FUS) splicing factors in IKBKAP splicing. To better evaluate the ExSpeU1s therapeutic efficacy, I transduced FD patient’s fibroblasts with a lentiviral vector expressing the most active ExSpeU1. This resulted in a complete rescue of the exon skipping defect and improvement in IKAP protein expression. Most importantly, intraperitoneal delivery of ExSpeU1s by AAV9 into a transgenic mouse model, that recapitulates the tissue-specific mis-splicing seen in FD patients, corrected the aberrant splicing patterns in several tissues increasing the amount of the corresponding IKAP protein. All together, these results identify novel regulatory splicing factors involved in the IKBKAP exon 20 regulation and provide the proof of principle that ExSpeU1s delivered in vivo by AAV vectors represent a novel therapeutic strategy for FD.
Keywords: Familial Dysautonomia, IKBKAP, IKAP, splicing, splicing defects, ExSpeU1, U1 snRNA, mouse model, AAV
Published in RUNG: 26.03.2018; Views: 3970; Downloads: 123
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Abstract: A strain of Klebsiella oxytoca DSM 29614 is grown on sodium citrate in the presence of 50 mg l−1 of Hg as Hg(NO3)2. During growth, the strain produces an extracellular polymeric substance (EPS), constituted by a mixture of proteins and a specific exopolysaccharide. The protein components, derived from the outer membrane of cells, are co-extracted with the extracellular exopolysaccharide using ethanol. The extracted EPS contains 7.5% of Hg (total amount). This indicates that EPS is an excellent material for the biosorption of Hg2+, through chemical complexation with the EPS components. The binding capacity of these species towards Hg2+ is studied by cyclic voltammetry, and Hg L3-edge XANES and EXAFS spectroscopy. The results found indicate that Hg2+ is mainly bound to the nitrogen of the imidazole ring or other N-heterocycle compounds. The hydroxyl moities of sugars and/or the carboxyl groups of two glucuronic acids in the polysaccharide can also play an important role in sequestring Hg2+ ions. However, N-heterocyclic groups of proteins bind Hg2+ faster than hydroxyl and carboxyl groups of the polysaccharide.
Keywords: Klebsiella oxytoca, Biosorption, Proteins, Polysaccharide, Cyclic voltammetry, EXAFS analysis
Published in RUNG: 03.03.2017; Views: 4134; Downloads: 0
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Abstract: Bacteria Klebsiella oxytoca (DSM 29614) secret a specific exopolysaccharide (EPS) in the presence of Fe(III)-citrate, as sole carbon and energy source, and produces after 7 days an iron gel precipitate (Fe-EPS) [1]. In the presence of palladium or other metal species, the culture of K. oxytoca can produce other mono or bimetal species (Pd-EPS, FePd-EPS) [2]. These bio-materials may be used as green biogenerated catalysts or for other biotechnological purposes. In this work we present the analysis of the morphology and the chemical state of the metals in mono and bi-metallic (Fe-EPS, Pd-EPS, FePd-EPS) complexes, by a combination of micro X-ray fluorescence, X-ray absorption spectroscopy methods (XANES and EXAFS), and transmission electron microscopy. The results show that iron in monometallic Fe-EPS and bimetalic FePd-EPS is in the mineralized form of iron oxides/hydroxides, predominantly in the form of Fe3+, with a small amount of Fe2+ in the structure, as a mixture of different nano-crystalline iron oxides and hydroxides. Palladium on the other hand is found as Pd(0) in the form of metallic nanoparticles with fcc structure in both, bi-metallic (FePd-EPS) and mono-metallic (Pd-EPS) species. In bi-metallic species Pd and Fe nanoparticles agglomerate in larger clusters, but they remain spatially separated. Access to the SR facilities at ESRF (beamline ID21, project LS-2225), DESY (beamline C) Hamburg (project I-20110511 EC) and ELETTRA (beamline XAFS, project 20115112) is acknowledged.
Keywords: Klebsiella oxytoca, Fe, Pd, XANES, EXAFS, exopolysaccharide, FePd-EPS
Published in RUNG: 28.06.2016; Views: 5114; Downloads: 0
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