STUDY OF BIOPROBE FOR DNA POLYMERASES

茄纳皮酮A是一种从真菌中分离出来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lambda(polβ和lambda)的活性。化合物的IC50值分别为Polβ30微米和POI lambda 37微米。因为POIβ和lambda属于一个家族，而且它们的三维结构被认为彼此高度相似，所以我们使用了polbeta。分析了与茄皮酮A的生化关系。在POIβ上，茄皮酮A与DNA模板和核苷酸底物发生拮抗竞争。BlAcore分析表明，茄那皮酮A选择性地与POIβ的N端8 kDa结构域结合。已知该结构域结合单链DNA，提供5-磷酸识别有间隙的DNA，并将糖-磷酸键3切割到完整的无嘌呤/脱嘧啶(AP)位置(即AP裂解酶活性)，包括5-脱氧核糖磷酸裂解酶活性。茄那皮酮A抑制单细胞…更多的e-链DNA结合活性，但不影响缝隙DNA结构中5-磷酸识别和AP裂解酶的活性。脱氢altenusin是一种哺乳动物DNA聚合酶α(pol.Alpha)，但不影响包括其他脊椎动物POL在内的其他复制DNA聚合酶的活性。阿尔法。在本研究中，我们提纯或合成了各种轻微修饰的脱氢青蒿素衍生物，并利用它们，研究了化学结构与抑制作用的关系，以及脱氢青蒿素的体内外作用，以确定其在多大程度上受到POL的影响。α活性抑制会影响细胞的增殖。脱氢阿糖胞苷使人胃癌细胞株NTJGC3细胞停滞于G1/S期，抑制细胞增殖，并阻止胸腺嘧啶核苷掺入细胞内，提示其通过抑制Pol而阻断体内DNA复制的第一步。阿尔法。较少

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