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Modified nucleotides and DNA for electrochemical labelling and defined display of small molecules
Krömer, Matouš ; Hocek, Michal (advisor) ; Křen, Vladimír (referee) ; Vrábel, Milan (referee)
This thesis is focused on enzymatic construction of DNA probes for electrochemical labelling, bioconjugations and, in the final part, building on knowledge gathered in previous chapters, it describes a method useful for construction of highly functionalized base-modified DNA enabling defined multivalent display of glycosides. In first chapter, a chemical route to diol-bearing nucleotides was found. Sonogashira reaction facilitated access to alkyne-tethered diols and subsequent catalytic hydrogenation, described for the first time in the literature, provided protection-free method for obtaining nucleotide diols tethered via flexible sp3 hybridized linker. Cleavage of alkane-linked, but not alkyne-linked, nucleotide diols yielded aliphatic nucleotide aldehyde. All nucleotides were found to be good substrates for KOD XL DNA polymerase in both primer extension and polymerase chain reaction, apart from aldehyde-linked dUCHO TP nucleotide, which performed well in PEX reaction, but gave PCR products only in a mixture with natural dTTP. This could be overcome by cleavage of diol-modified DNA, which also yielded aldehyde-functionalized dsDNA. All reactive probes were examined for bioconjugations with fluorescent hydrazine, reductive amination with lysine or lysine-containing peptides or other molecules...
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New cross-coupling reactions and C-H activations for synthesis of modified nucleobase analogues
Krömer, Matouš ; Hocek, Michal (advisor) ; Kotora, Martin (referee)
6 Abstract A novel methodology of construction of 6,7-disubstituted 7-deazapurines by serial of orthogonal cross-coupling reactions was developed. The Liebeskind-Srogl reaction was optimized and the reactivity of a few boronic acids was explored. Library of 3×3 disubstuted analogues of deazapurines was synthesized utilizing this method. In the second part of this work, the scope of direct alkylation of purine derivatives was verified. Keywords purine, 7-deazapurine, orthogonal cross-coupling reactions, C-H activations
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