5-(Hydroxymethyl)-2-furfural: A selective inhibitor of DNA polymerase λ and terminal deoxynucleotidyltransferase

5-(Hydroxymethyl)-2-furfural (HMF), a pyrolysate of carbohydrate isolated from instant coffee (Coffea arabica L.), selectively inhibits the activities of mammalian DNA polymerase λ (pol λ) and terminal deoxynucleotidyltransferase (TdT) which are family X pols, in vitro. The compound influenced neither the activities of replicative DNA polymerases such as α, δ, and ε, nor even the activity of pol β which is from the same family and thought to have a very similar three-dimensional structure to the pol β-like region of pol λ. Since parts of HMF such as furan, furfuryl alcohol, and 2-furaldehyde did not influence the activities of any enzymes tested, the substituted form of furan with a hyroxymethyl group and a formyl group might be important for the inhibition of pol λ and TdT. The inhibitory effect of HMF on intact pol λ (i.e., residues 1–575), a truncated pol λ lacking the N-terminal BRCA1 C-terminus domain (133–575, del-1 pol λ) and another truncated pol λ lacking the N-terminal proline-rich region (245–575, del-2 pol λ) was dose-dependent, and 50% inhibition was observed at a concentration of 26.1, 10.3, and 4.6 μM, respectively. The IC50 value of HMF for TdT was the same as that for del-2 pol λ (5.5 μM). The HMF-induced inhibition of both pol λ and TdT activities was competitive with respect to both the DNA template-primer and the dNTP substrate. On the basis of these results, HMF was suggested to bind to the pol β-like region of pol λ and TdT.