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Codon usage and isoacceptor tRNA expression as a mechanism to control gene expression
Daumová, Lenka ; Čáp, Michal (advisor) ; Štěpánek, Luděk (referee)
Genetic code is defined as a set of rules, which encode aminoacid sequences in proteins, according to codon usage. It is widely known, that there are multiple codons for most aminoacids, thanks to the degeneracy of the genetic code. There was a hypothesis, that silent mutations, which result in a synonymous codon and not in incorporating of a different aminoacid into the peptide chain, don't affect the gene expression. Later however, it was found through more detailed research on molecular level, that codon usage bias is in fact one of the factors, that regulate translation effectivity and rate, mRNA stability or even protein folding and gene expression. There has been many studies published on these topics. This bachelor thesis is a review of these studies. First, I summarize basic information on tRNA, its structure and modifications in anticodon loop. Next I write about base pairing between codon and anticodon, including the non-canonical wobble base pairing. Then I emphasize on codon bias, its causes, its relationship with genome GC content. I also include some author's conjectures about how to approach this phenomenon, which codons are optimal and what is the impact of codon usage bias on translation efficiency. I cite many studies on this topis, which was researched on many model organisms,...
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A study of the HCV IRES variability: An experimental approach coupled with design of a large-scale mutation database
Khawaja, Anas Ahmad ; Pospíšek, Martin (advisor) ; Hirsch, Ivan (referee) ; Valášek, Leoš (referee)
Translation initiation in the hepatitis C virus (HCV) occurs through a cap- independent mechanism that involves an internal ribosome entry site (IRES) capable of interaction with and utilization of the eukaryotic translational machinery. We focused on the structural configuration of the different HCV-IRES domains and the impact of IRES primary sequence variations on secondary structure conservation and function. For this purpose we introduced into our laboratory, methods such as denaturing gradient and temperature gradient gel electrophoresis for screening the degree of heterogeneity and total amount of HCV-IRES variability accumulated in HCV infected patients over a period of time. The selected samples showed variable migration pattern of the HCV-IRES (from all the patients) visualized in DGGE and TGGE, were sequenced and evaluated for translation efficiency using flow cytometry. In some cases, we discovered that multiple mutations, even those scattered across different domains of HCV-IRES, led to restoration of the HCV-IRES translational activity, although the individual occurrences of these mutations were found to be deleterious. We propose that such observation may be attributed to probable long- range inter- and/or intra-domain functional interactions. We established a large-scale HCV-IRES...
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