Modifikace DNA reaktivními skupinami pro biokonjugace a cross-linkování s peptidy a proteiny obsahujícími arginin

Leone, Denise Liu

This PhD thesis describes the development and the synthesis of DNA-reactive probes bearing 1,3- diketone or phenylglyoxal moieties which can be used for cross-linking with arginine-containing peptides or proteins. The general strategy was based on the functionalization of the position 5 of pyrimidines via aqueous Sonogashira coupling or click reaction (CuAAC) with reactive building blocks to synthesize modified 2'-deoxyribonucleoside monophosphates and triphosphates (dNMPs/dNTPs). The following step was the enzymatic incorporation of such functionalized dNTPs into DNA via primer extension (PEX). In parallel, the monophosphate derivatives were used as model compounds for reactions with arginine and arginine-containing peptides to test the reactivity of the probes. For the reactive candidates, model reactions on the modified-DNA with arginine or Arg-containing peptides were performed and the DNA-probes were used in cross- linking reactions with Arg-rich DNA-binding proteins. The development of such probes was not straightforward but required the investigation of diverse candidates. More importantly the balance between reactivity and stability of the reactive group was the focus point and the challenge in the design of the reactive building blocks. The first candidate consisted of thiazolidine masked...

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