Biotin‐ and Chloroalkane‐Modified Nucleotides for Dual Display of Proteins on DNA Through Streptavidin and Halotag Binding

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Abstract
We designed and synthesized 2’‐deoxyribonucleoside triphosphates (dNTPs) bearing chlorohexyl and biotin modifications attached to position 7 of 7‐deazaadenine or to position 5 of cytosine. The modified dNTPs served as substrates for KOD XL DNA polymerase in the enzymatic construction of DNA bearing either one or a combination of the two modifications. The chlorohexyl‐linked DNA underwent covalent reactions with halotag protein, while the biotin‐linked DNA formed stable non‐covalent complexes with monomeric streptavidin. The dually modified DNA bearing both modifications at different distances apart from each other were used for sequence‐specific, spatially defined display of both halotag and streptavidin proteins. The study provides the foundation for display of proteins on DNA rulers.
Funding Information
  • Grantová Agentura České Republiky (25–15592S)

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