63(P-37) 糸状菌の生産するDNAポリメラーゼβおよびλ選択的阻害剤、solanapyrone A(ポスター発表の部)

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Mammalian cells contain at least twelve types of DNA polymerase (pol. α, β, γ, δ, ε, ζ, η, θ, ι, λ, κ, and μ) We screened for natural compounds that selectively inhibit pol. β or λ. The purpose of this screenings was to use the compounds as tools or molecular probes to distinguish these DNA polymerases (pols) and to clarify both biological and in vivo function. Pol. β consists of N-terminal 8kDa and C-terminal 31kDa domains. The N-terminal domain was originally characterized as a single-stranded DNA (ssDNA) binding domain. It specifically binds the 5'-phosphate in gapped DNA. Pol. β catalyzes removal of deoxyribose phosphate (dRP) from apurinic-apyrimidinic (AP) endonuclease incised AP site via β-elimination. This dRP lyase activity resides in the 8kDa domain of pol. β. Although the in vivo function of the family enzyme, pol. λ, is unclear yet, it appears to play in the similar situation to pol. β. We have analyzed the structure and function of both pol. β and pol. λ, using the inhibitor from two different points of view; to understand the precise role of each of the pols in vivo, and to develop a drug design strategy for cancer chemotherapy agents. Since pol. β is an essential enzyme for nucleotide excision repair, the pol. β inhibitor may lead to block the DNA repair systems for rescue cancer cells under the radiation-therapy or chemotherapy clinically. A compound, which selectively inhibits the activities of mammalian pols β and λ in vitro, was found from a fungus (SUT01B-1-2). The agent was identified solanapyrone A by spectrometric analysis. Solanapyrone A is a plant phytotoxin isolated together with other toxic metabolites from Alternaria solani, causal fungus of early blight disease of potato. The IC_<50> values of this compound were 30μM for pol. β and 37μM for pol. λ. No pol. λ inhibitors have been reported yet. Since pols β and λ are in a family and their three-dimensional structures are considered to be highly similar to each other, we used pol. β to analyze the biochemical relationship with solanapyrone A. On pol. β, solanapyrone A antagonistically competed with both the DNA template and the nucleotide substrate. BIAcore analysis demonstrated that solanapyrone A bound selectively to the DNA template-binding domain (8kDa domain) of pol. β. From the facts that the domain has the activities of ssDNA binding, 5'-phosphate recognition of gapped DNA, and AP lyase activity, solanapyrone A was found to inhibit the ssDNA binding activity.
天然有機化合物討論会の論文
2002-09-01

63(P-37) 糸状菌の生産するDNAポリメラーゼβおよびλ選択的阻害剤、solanapyrone A(ポスター発表の部)

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