Direct Conjugation of Peptides and 5‑Hydroxymethylcytosine
in DNA
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Abstract
Recent
discovery of functional 5-hydroxymethylcytosine in vertebrate
genomes prompted for elaboration of methods to localize this modification
at the nucleotide resolution level. Among several covalent modification-based
approaches, atypical activity of cytosine-5 DNA methyltransferases
to couple small molecules to 5-hydroxymethylcytosine stands out for
acceptance of broad range of ligands. We went further to explore the
possibility for methyltransferase-maintained coupling of compounds
possessing autonomous functions. Functionalization of DNA was achieved
by direct conjugation of chemically synthesized peptides of regular
structure. Sequence, residue, and position-specific coupling of DNA
containing 5-hydroxymethylcytosine and different peptides has
been demonstrated, with the nature of the resulting conjugates confirmed
by protease treatment and mass spectrometry. Coupling products were
compatible with affinity-driven separation from the unmodified DNA.
This approach highlights an emerging avenue toward the enzymatic,
sequence-specific DNA functionalization, enabling a single step merge
of the DNA and peptide moieties into a bifunctional entity