Development of separation and detection of functional life molecules using combinatorial chemistry and ribozyme system

利用组合化学和核酶系统开发功能生命分子的分离和检测

• 批准号: 09440253
• 负责人: SUGIMOTO Naoki
• 金额: $ 7.87万
• 依托单位: Konan University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B).
• 财政年份: 1997
• 资助国家: 日本
• 起止时间: 1997 至 2000
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-09440253/
• 关键词: Combinatorial chemistry; Ribozyme; Deoxyribozyme; RNA cleavage; Calcium ion; Metal ion; DNA chip; Avidin- biotin interaction; リポザイム; RNA切断活性; DNA; RNA; 反応速度; 分子認識; ペプチド; コンビナトリアルケミストリー; インビトロセレクション; RecA; 機能性材料; ファージディスプレイ; 遺伝子診断; 核酸

Catalytic DNA is a promising class of nucleic acid enzyme for possible use as a therapeutic agent and is also interesting in comparison with catalytic RNAs. In this study, we investigated the effect of metal ions and sequence on an original deoxyribozyme, d(GCCTGGCAG_1 G_2 C_3 T_4 A_5 G_6 C_7 T_8 A_9 C_<10> A_<11> A_<12> C_<13> G_<14> A_<15> GTCCCT), which binds to an RNA substrate, r(GAAGACA↓UGCCAGCG), cleaving the RNA substrate at one site indicated by the arrow. The results show that the ability of metal ions to promote the RNA cleavage reaction by the original deoxyribozyme is Mn^<2+> >Mg^<2+> >Ca^<2+> * Ba^<2+>. This result is very similar to the previous tendency observed in the case of a hammerhead ribozyme. Thus, these results suggest that RNA cleavage by the deoxyribozyme dependence on metal ions for catalysis is similar to that with ribozymes. On the other hand, a nucleotide deletion from the active domain of the original deoxyribozyme results in a novel deoxyribozyme with high Ca^<2+> -dependency, a situation which is not observed with the original deoxyribozyme or hammerhead ribozymes.

催化DNA是一类有前途的核酸酶，可能用作治疗剂，并且与催化RNA相比也很有趣。在这项研究中，我们研究了金属离子和序列对原始脱氧核酶d（GCCTGGCAG_1 G_2 C_3 T_4 A_5 G_6 C_7 T_8 A_9 C_<10>A<11>_<12>C_<13>G_<14>A_<15>GTCCCT）的影响，该核酶与RNA底物r（GAAGACA↓UGCCAGCG）结合，在箭头所示的一个位点切割RNA底物。结果表明，金属离子对原脱氧核酶切割RNA的促进作用大小依次为Mn^2+&gt;Mg^2+&gt;Ca ^2+* Ba^2+。这一结果与先前在锤头状核酶的情况下观察到的趋势非常相似。因此，这些结果表明，RNA切割的脱氧核酶依赖于金属离子的催化是类似的核酶。另一方面，从原始脱氧核酶的活性结构域中删除一个核苷酸会产生一种具有高度Ca^2+依赖性的新型脱氧核酶，这种情况在原始脱氧核酶或锤头状核酶中没有观察到。

项目成果

N.Sugimoto: "α-β structural Transition of Short Oligopeptides by Water/Organic Solvent Titration"Chem.Lett.. 637-638 (1999)

N.Sugimoto：“通过水/有机​​溶剂滴定法实现短寡肽的 α-β 结构转变”Chem.Lett.. 637-638 (1999)

J.Kawakami, H.Imanaka, Y.Yokota, and N.Sugimoto: "In Vitro Selection of Aptamers That Act with Zn2+"J.Inorg.Biochem. 82. 197-206 (2000)

J.Kawakami、H.Imanaka、Y.Yokota 和 N.Sugimoto：“与 Zn2 作用的适体的体外选择”J.Inorg.Biochem。

Peng Wu: "Transition characteristics and thermodynamic analysis of DNA duplex formation : a quantitative consideration for the extent of duplex association"Nucleic Acids Res.. 28. 4762-4768 (2000)

吴鹏：“DNA双链体形成的转变特征和热力学分析：双链体缔合程度的定量考虑”《核酸研究》28. 4762-4768 (2000)

N.Sugimoto, M.Nakano, and S.Nakano: "Thermodynamics-Structure Relationship of Single Mismatches in RNA/DNA Duplexes"Biochemistry. 39. 11270-11281 (2000)

N.Sugimoto、M.Nakano 和 S.Nakano：“RNA/DNA 双链体中单个错配的热力学-结构关系”生物化学。

T.Ohmichi, H.Nakamuta, K.Yasuda, and N.Sugimoto: "Kinetic Property of Bulged Helix Formation : Analysis of Kinetic Behavior Using Nearest-Neighbor Parameters"J.Am.Chem.Soc.. 122. 11286-11294 (2000)

T.Ohmichi、H.Nakamuta、K.Yasuda 和 N.Sugimoto：“凸出螺旋形成的动力学特性：使用最近邻参数分析动力学行为”J.Am.Chem.Soc.. 122. 11286-11294 (