DNA with a 2-pyridyldithio group at the C2' atom: a promising tool for the crosslinking of the MutS protein preserving  its functional activity

Perry, S.A.; Kubareva, E.A.; Monakhova, M.V.; Trikin, R.M.; Kosaretskiy, E.M.; Romanova, E.A.; Metelev, V.G.; Friedhoff, P.; Oretskaya, T.S.

Авторы: Perry S.A., Kubareva E.A., Monakhova M.V., Trikin R.M., Kosaretskiy E.M., Romanova E.A., Metelev V.G., Friedhoff P., Oretskaya T.S.
Журнал: Russian Journal of Bioorganic Chemistry
Том: 47
Номер: 2
Год издания: 2021
Издательство: Maik Nauka/Interperiodica Publishing
Местоположение издательства: Russian Federation
Первая страница: 446
Последняя страница: 459
Оригинальные версии: ДНК с 2-пиридилдитиогруппой при С2'-атоме – перспективные инструменты для фиксации белка MutS с сохранением его функциональной активности
DOI: 10.1134/S1068162021020205
Аннотация: Noncovalent interactions between DNA and proteins are vital for cells. DNA-protein crosslinking may occur due to various endogenous and exogenous factors, as well as chemotherapeutic agents, and often affects the performance of DNA-binding proteins. Oligodeoxyribonucleotides with a reactive group can serve as a tool to investigate the structure and action of DNA-binding proteins. Here, we report the crosslinking of MutS, which is a sensor protein of DNA mismatch repair (MMR) in Escherichia coli, via the protein’s cysteine. This was realized via DNA that harboured a 2′-deoxy-2′-[3-(2-pyridyldithio)propionamide] group as part of a nucleoside at a given position, and two fluorophores. The proposed DNA duplex consisted of complementary oligonucleotides with breaks in the top and bottom strands. This feature allowed us to introduce the reactive group at different positions and made it possible to synthesize only the central fragment of the DNA with a minimal length. The MutS-DNA conjugate was obtained by thiol-disulfide exchange with quantitative yield and separated from the unreacted DNA by size-exclusion chromatography. Fluorescence resonance energy transfer was used to show that MutS remained functionally active within the conjugate, since the protein was able to change its conformation and DNA conformation as well.
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DNA with a 2-pyridyldithio group at the C2' atom: a promising tool for the crosslinking of the MutS protein preserving its functional activityстатья

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