STUDY OF BIOPROBE FOR DNA POLYMERASES

DNA聚合酶生物探针的研究

• 批准号: 12660103
• 负责人: FUMIO SUGAWARA
• 金额: $ 2.5万
• 依托单位: TOKYO UNIVERSITY OF SCIENCE
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2000
• 资助国家: 日本
• 起止时间: 2000 至 2001
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-12660103/
• 关键词: sanapyrone A; DNA polvmerase beta and lambda; phytotoxin; dehydroaltenusin; DNA replication; NMR; solanapyrone; lithochol酸; ネルボン酸; ヒト免疫不全症候群ウィルス; HIV-1RT

Solanapyrone A, a phytotoxin and enzyme inhibitor isolated from a fungus, electively inhibits the activities of mammalian DNA polymerase beta and lambda (pol beta and lambda) in vitro. The IC50 values of the compound were 30 microm for pol beta and 37 microm for poi lambda. Because poi beta and lambda are in a family and their three-dimensional structures are thought to be highly similar to each other, we used pol beta to. analyze the biochemical relationship with solanapyrone A. On poi beta, solanapyrone A antagonistically competed with both the DNA template and the nucleotide substrate. BlAcore analysis demonstrated that solanapyrone A bound selectively to the N-terminal 8-kDa domain of poi beta. This domain is known to bind single-stranded DNA, provide 5 -phosphate recognition of gapped DNA, and cleave the sugar-phosphate bond 3 to an intact apurinic/apyrimidinic (AP) site (i. e. AP lyase activity) including 5 -deoxyribose phosphate lyase activity. Solanapyrone A inhibited the singl … More e-stranded DNA-binding activity but did not influence the activities of the 5 -phosphate recognition in gapped DNA structures and the AP lyase. Dehydroaltenusin was found to be an inhibitor of mammalian DNA polymerase alpha (pol. alpha) in vitro, but did not influence the activities of the other replicative DNA polymerases including even other vertebrate pol. alpha. In this study, we purified or synthesized various slightly modified derivatives of dehydroaltenusin, and using them, investigated the relationship between the chemical structure and the inhibitory effects, and the in vitro and in vivo effects of dehydroaltenusin to determine to what extent the pol. alpha activity inhibition influences cell proliferation. Dehydroaltenusin inhibited the cell proliferation of the human gastric cancer cell line NTJGC-3 by arresting the cells at G1/S-phase, and prevented the incorporation of thymidine into the cells, indicating that it blocks the primary step of in vivo DNA replication by inhibiting pol. alpha. Less

茄吡喃酮A是从真菌中分离的植物毒素和酶抑制剂，在体外选择性地抑制哺乳动物DNA聚合酶β和λ（pol β和λ）的活性。该化合物的IC 50值对pol β为30 μ m，对pol λ为37 μ m。因为poi beta和lambda是一个家族，并且它们的三维结构被认为彼此高度相似，所以我们使用poi beta来。分析与茄吡喃酮A的生化关系。在poi β上，茄吡喃酮A拮抗地与DNA模板和核苷酸底物竞争。BlAcore分析表明，茄吡喃酮A选择性地结合到poi β的N-末端8-kDa结构域。已知该结构域结合单链DNA，提供缺口DNA的5 -磷酸识别，并将糖-磷酸键3切割至完整的脱嘌呤/脱嘧啶（AP）位点（即，e. AP裂解酶活性），包括5 -脱氧核糖磷酸裂解酶活性。茄尼吡喃酮A抑制单细胞凋亡， ...更多信息 e-链DNA结合活性，但不影响缺口DNA结构中的5 -磷酸识别和AP裂解酶的活性。发现脱氢阿替努辛在体外是哺乳动物DNA聚合酶α（pol. alpha）的抑制剂，但不影响其它复制性DNA聚合酶的活性，包括甚至其它脊椎动物pol. alpha。在这项研究中，我们纯化或合成了各种轻微修饰的脱氢Altenusin衍生物，并使用它们，研究化学结构和抑制作用之间的关系，以及脱氢Altenusin的体外和体内作用，以确定在多大程度上pol. alpha活性抑制影响细胞增殖。脱氢阿替努辛通过将细胞阻滞在G1/S期来抑制人胃癌细胞系NTJGC-3的细胞增殖，并阻止胸苷掺入细胞，表明其通过抑制pol α阻断体内DNA复制的初级步骤。少

项目成果

F.SUGAWARA: "Japanese research and developments on plant pathogens as weed control agent"J. Crop Prot.. 4. 277-285 (2001)

F.SUGAWARA：“日本对植物病原体作为杂草控制剂的研究和开发”J。

Y.MLZUSHINA, N.KASAI, F.SUGAWARA, A.IIDA, H.YOSHIDA, K.SAKAGUCHI.: "Three-dimensional structural model analysis of the binding site of lithocholic acid, an inhibitor of DNA polymerase β and DNA topoisomerase II"J. Biochem. 130. 657-664 (2001)

Y.MLZUSHINA、N.KASAI、F.SUGAWARA、A.IIDA、H.YOSHIDA、K.SAKAGUCHI.：“DNA 聚合酶 β 和 DNA 拓扑异构酶 II 抑制剂石胆酸结合位点的三维结构模型分析“生物化学杂志。130。657-664（2001）

H.SAHARA, S.HANASHIMA, T.YAMAZAKI, S.TAKAHASHI, F.SUGAWARA, S.OHTANI, M.ISHIKAWA, Y.MIZUSHINA, K.OHTA, N.TAKAHASHI, K.SHIMOZAWA, S.GASA, M.MORI, K.JIMBOW, K.SAKAGUCHI, N.SATO: "Anti-tumor effect of chemically synthesized sulpholipids deduced from sea urch

H.SAHARA、S.HANASHIMA、T.YAMAZAKI、S.TAKAHASHI、F.SUGAWARA、S.OHTANI、M.ISHIKAWA、Y.MIZUSHINA、K.OHTA、N.TAKAHASHI、K.SHIMOZAWA、S.GASA、M.

Y.MIZUSHINA, Y.SUGIYAMA, H.YOSHIDA, S.HANASHIMA, T.YAMAZAKI, S.KAMISUKI, K.OHTA, M.TAKEMURA, T.YAMAGUCHI, A.MATSUKAGE, S.YOSHIDA, M.SANEYOSHI, F.SUGAWARA, S.SAKAGUCHI: "Galactosyldiacylglycerol, a mammalian DNA polymerase α-specific inhibitoi form sea alg

Y.MIZUSHINA、Y.SUGIYAMA、H.YOSHIDA、S.HANASHIMA、T.YAMAZAKI、S.KAMISUKI、K.OHTA、M.TAKEMURA、T.YAMAGUCHI、A.Matsukage、S.YOSHIDA、M.Saneyoshi、F. SUGAWARA, S.SAKAGUCHI：“半乳糖基二酰基甘油，一种哺乳动物 DNA 聚合酶 α 特异性抑制剂，形成海藻。

C.Murakami, M.Takemura, Y.Sugiyama, S.Kamisuki, H.Asahara, M.Kawasaki, T.Ishidoh, S.Linn, S.Yoshida, F.Sugawara, H.Yoshida, K.Sakaguchi, Y.Mizushina: "Vitamin A-related compounds, all-trans retinal and retinoic acids, selectively inhibit activities of mam

C.Murakami、M.Takemura、Y.Sugiyama、S.Kamisuki、H.Asahara、M.Kawasaki、T.Ishidoh、S.Linn、S.Yoshida、F.Sugara、H.Yoshida、K.Sakaguchi、Y.