The Novel Chemical Mechanism of the Twister Ribozyme

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• 作者：Timothy J. Wilson ; Yijin Liu ; Christof Domnick ; Stephanie Kath-Schorr ; David M. J. Lilley
• 刊名：Journal of the American Chemical Society
• 出版年：2016
• 出版时间：May 18, 2016
• 年：2016
• 卷：138
• 期：19
• 页码：6151-6162
• 全文大小：627K
• 年卷期：0
• ISSN：1520-5126

文摘

We describe the multifactorial origins of catalysis by the twister ribozyme. We provide evidence that the adenine immediately 3′ to the scissile phosphate (A1) acts as a general acid. Substitution of ring nitrogen atoms indicates that very unusually the N3 of A1 is the proton donor to the oxyanion leaving group. A1 is accommodated in a specific binding pocket that raises its pK_a toward neutrality, juxtaposes its N3 with the O5′ to be protonated, and helps create the in-line trajectory required for nucleophilic attack. A1 performs general acid catalysis while G33 acts as a general base. A 100-fold stereospecific phosphorothioate effect at the scissile phosphate is consistent with a significant stabilization of the transition state by the ribozyme, and functional group substitution at G33 indicates that its exocyclic N2 interacts directly with the scissile phosphate. A model of the ribozyme active site is proposed that accommodates these catalytic strategies.