Efficient and Rapid Template-Directed Nucleic Acid Copying Using 2′-Amino-2′,3′-dideoxyribonucleoside−5′-Phosphorimidazolide Monomers

• Published in: Journal of the American Chemical Society 2009-10, Vol.131 (40), p.14560-14570
• Main Authors: Schrum, Jason P, Ricardo, Alonso, Krishnamurthy, Mathangi, Blain, J. Craig, Szostak, Jack W
• Format: Article
• Language: English
• Subjects: Dideoxynucleosides - chemistry; Dideoxynucleosides - metabolism; DNA - biosynthesis; DNA - chemical synthesis; DNA - genetics; Imidazoles - chemistry; Imidazoles - metabolism; Nucleic Acids - biosynthesis; Nucleic Acids - chemical synthesis; Nucleic Acids - genetics; RNA - biosynthesis; RNA - chemical synthesis; RNA - genetics; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Templates, Genetic
• ISSN: 0002-7863; 1520-5126
• DOI: 10.1021/ja906557v

The development of a sequence-general nucleic acid copying system is an essential step in the assembly of a synthetic protocell, an autonomously replicating spatially localized chemical system capable of spontaneous Darwinian evolution. Previously described nonenzymatic template-copying experiments have validated the concept of nonenzymatic replication, but have not yet achieved robust, sequence-general polynucleotide replication. The 5′-phosphorimidazolides of the 2′-amino-2′,3′-dideoxyribonucleotides are attractive as potential monomers for such a system because they polymerize by forming 2′→5′ linkages, which are favored in nonenzymatic polymerization reactions using similarly activated ribonucleotides on RNA templates. Furthermore, the 5′-activated 2′-amino nucleotides do not cyclize. We recently described the rapid and efficient nonenzymatic copying of a DNA homopolymer template (dC15) encapsulated within fatty acid vesicles using 2′-amino-2′,3′-dideoxyguanosine−5′-phosphorimidazolide as the activated monomer. However, to realize a true Darwinian system, the template-copying chemistry must be able to copy most sequences and their complements to allow for the transmission of information from generation to generation. Here, we describe the copying of a series of nucleic acid templates using 2′-amino-2′,3′-dideoxynucleotide−5′-phosphorimidazolides. Polymerization reactions proceed rapidly to completion on short homopolymer RNA and LNA templates, which favor an A-type duplex geometry. We show that more efficiently copied sequences are generated by replacing the adenine nucleobase with diaminopurine, and uracil with C5-(1-propynyl)uracil. Finally, we explore the copying of longer, mixed-sequence RNA templates to assess the sequence-general copying ability of 2′-amino-2′,3′-dideoxynucleoside−5′-phosphorimidazolides. Our results are a significant step forward in the realization of a self-replicating genetic polymer compatible with protocell template copying and suggest that N2′→P5′-phosphoramidate DNA may have the potential to function as a self-replicating system.