DNA catalysts with tyrosine kinase activity

We show that DNA catalysts (deoxyribozymes, DNA enzymes) can phosphorylate tyrosine residues of peptides. Using in vitro selection, we identified deoxyribozymes that transfer the γ-phosphoryl group from a 5′-triphosphorylated donor (a pppRNA oligonucleotide or GTP) to the tyrosine hydroxyl acceptor of a tethered hexapeptide. Tyrosine kinase deoxyribozymes that use pppRNA were identified from each of N₃₀, N₄₀, and N₅₀ random-sequence pools. Each deoxyribozyme requires Zn²⁺, and most additionally require Mn²⁺. The deoxyribozymes have little or no selectivity for the amino acid identities near the tyrosine, but they are highly selective for phosphorylating tyrosine rather than serine. Analogous GTP-dependent DNA catalysts were identified and found to have apparent K_m(GTP) as low as ∼20 μM. These findings establish that DNA has the fundamental catalytic ability to phosphorylate the tyrosine side chain of a peptide substrate.