Efforts toward Expansion of the Genetic Alphabet: Structure and Replication of Unnatural Base Pairs

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• 作者：Shigeo Matsuda ; Jeremiah D. Fillo ; Allison A. Henry ; Priyamrada Rai ; Steven J. Wilkens ; Tammy J. Dwyer ; Bernhard H. Geierstanger ; David E. Wemmer ; Peter G. Schultz ; Glen Spraggon ; Floyd E. Romesberg
• 刊名：Journal of the American Chemical Society
• 出版年：2007
• 出版时间：August 29, 2007
• 年：2007
• 卷：129
• 期：34
• 页码：10466 - 10473
• 全文大小：552K
• 年卷期：v.129,no.34(August 29, 2007)
• ISSN：1520-5126

文摘

Expansion of the genetic alphabet has been a long-time goal of chemical biology. A third DNAbase pair that is stable and replicable would have a great number of practical applications and would alsolay the foundation for a semisynthetic organism. We have reported that DNA base pairs formed betweendeoxyribonucleotides with large aromatic, predominantly hydrophobic nucleobase analogues, such aspropynylisocarbostyril (dPICS), are stable and efficiently synthesized by DNA polymerases. However, onceincorporated into the primer, these analogues inhibit continued primer elongation. More recently, we havefound that DNA base pairs formed between nucleobase analogues that have minimal aromatic surfacearea in addition to little or no hydrogen-bonding potential, such as 3-fluorobenzene (d3FB), are synthesizedand extended by DNA polymerases with greatly increased efficiency. Here we show that the rate of synthesisand extension of the self-pair formed between two d3FB analogues is sufficient for in vitro DNA replication.To better understand the origins of efficient replication, we examined the structure of DNA duplexescontaining either the d3FB or dPICS self-pairs. We find that the large aromatic rings of dPICS pair in anintercalative manner within duplex DNA, while the d3FB nucleobases interact in an edge-on manner, muchcloser in structure to natural base pairs. We also synthesized duplexes containing the 5-methyl-substitutedderivatives of d3FB (d5Me3FB) paired opposite d3FB or the unsubstituted analogue (dBEN). In all, thedata suggest that the structure, electrostatics, and dynamics can all contribute to the extension of unnaturalprimer termini. The results also help explain the replication properties of many previously examined unnaturalbase pairs and should help design unnatural base pairs that are better replicated.