Eisenführ Alexander, Arora Paramjit S, Sengle Gerhard, Takaoka Leo R, Nowick James S, Famulok Michael
Kekulé-Institut für Organische Chemie und Biochemie, Rheinische Friedrich-Wilhelms-Universität Bonn, Gerhard-Domagk-Strasse 1, 53121 Bonn, Germany.
Bioorg Med Chem. 2003 Jan 17;11(2):235-49. doi: 10.1016/s0968-0896(02)00311-5.
The ability to generate RNA molecules that can catalyze complex organic transformations not only facilitates the reconstruction and plausibility of possible prebiotic reaction pathways but is also crucial for elucidating the potential of the application of RNA catalysts in organic syntheses. Iterative RNA selection previously identified a ribozyme that catalyzes the Michael addition of a cysteine thiol to an alpha,beta-unsaturated amide. This reaction is chemically similar to the rate limiting step of the thymidylate synthase reaction, which is the corresponding reaction of a cysteine thiol to the double-bond of the uracil nucleobase. Here we provide a detailed description of the synthesis of the ribozyme substrates and the substrate oligonucleotides used for its characterization and the investigation of the background reaction. We also describe the further characterization of the ribozyme with respect to substrate specificity. We show that the thiol group of the cysteine nucleophile is essential for the reaction to proceed. When substituted for a thiomethyl group, no reaction takes place.
生成能够催化复杂有机转化的RNA分子的能力，不仅有助于重建可能的前生物反应途径并使其具有合理性，对于阐明RNA催化剂在有机合成中的应用潜力也至关重要。此前通过迭代RNA筛选鉴定出一种核酶，它能催化半胱氨酸硫醇对α,β-不饱和酰胺的迈克尔加成反应。该反应在化学上类似于胸苷酸合成酶反应的限速步骤，即半胱氨酸硫醇与尿嘧啶核苷酸碱基双键的相应反应。在此，我们详细描述了用于其表征和背景反应研究的核酶底物及底物寡核苷酸的合成。我们还描述了核酶在底物特异性方面的进一步表征。我们表明，半胱氨酸亲核试剂的硫醇基团对于反应的进行至关重要。当被硫甲基取代时，不发生反应。
