Flavonoid glycoside: a new inhibitor of eukaryotic DNA polymerase α and a new carrier for inhibitor-affinity chromatography

Abstract

Two flavonoid glycosides, kaempferol 3- O-(6″-acetyl)-β-glucopyranoside (KAG) and quercetin 3- O-(6″-acetyl)-β-glucopyranoside (QAG), were found to be inhibitors of eukaryotic DNA polymerases from a Japanese vegetable, Petasites japonicus. These compounds inhibited the activities of mammalian replicative DNA polymerases (i.e., pol α, δ, and ε), but not other pol β, η, κ, and λ activities. KAG was a stronger inhibitor and more selective to pol α than QAG. The IC 50 values of KAG for pol α, δ, and ε were 41, 164, and 127 μM, respectively. The pol α inhibition by KAG was non-competitive with respect to both the DNA template-primer and the dNTP substrate. KAG and QAG did not influence the activities of prokaryotic DNA polymerases or other mammalian DNA metabolic enzymes such as human immunodeficiency virus type 1 reverse transcriptase, human telomerase, human DNA topoisomerase I and II, T7 RNA polymerase, and bovine deoxyribonuclease I. Therefore, we concluded that these flavonoid glycosides are moderate replicative DNA polymerase inhibitors leaning more relatively to pol α, and could be used as chromatographic carriers to purify the DNA polymerases rather than cytotoxic agents. We then made a KAG-conjugated column such as the epoxy-activated Sepharose 6B. In the column, pol α was selectively adsorbed and eluted.