Dataset Information

Enhanced Nonenzymatic RNA Copying with 2-Aminoimidazole Activated Nucleotides.

ABSTRACT: Achieving efficient nonenzymatic replication of RNA is an important step toward the synthesis of self-replicating protocells that may mimic early forms of life. Despite recent progress, the nonenzymatic copying of templates containing mixed sequences remains slow and inefficient. Here we demonstrate that activating nucleotides with 2-aminoimidazole results in superior reaction kinetics and improved yields of primer extension reaction products. This new leaving group significantly accelerates monomer addition as well as trimer-assisted RNA primer extension, allowing efficient copying of a variety of short RNA templates with mixed sequences.

SUBMITTER: Li L

PROVIDER: S-EPMC6326525 | biostudies-literature | 2017 Feb

REPOSITORIES: biostudies-literature

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Publications

Enhanced Nonenzymatic RNA Copying with 2-Aminoimidazole Activated Nucleotides.

Li Li L Prywes Noam N Tam Chun Pong CP O'Flaherty Derek K DK Lelyveld Victor S VS Izgu Enver Cagri EC Pal Ayan A Szostak Jack W JW

Journal of the American Chemical Society 20170124 5

Achieving efficient nonenzymatic replication of RNA is an important step toward the synthesis of self-replicating protocells that may mimic early forms of life. Despite recent progress, the nonenzymatic copying of templates containing mixed sequences remains slow and inefficient. Here we demonstrate that activating nucleotides with 2-aminoimidazole results in superior reaction kinetics and improved yields of primer extension reaction products. This new leaving group significantly accelerates mon ...[more]

PMID: 28117989

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Project description:Nonenzymatic RNA primer extension by activated mononucleotides has long served as a model for the study of prebiotic RNA copying. We have recently shown that the rate of primer extension is greatly enhanced by the formation of an imidazolium-bridged dinucleotide between the incoming monomer and a second, downstream activated monomer. However, the rate of primer extension is further enhanced if the downstream monomer is replaced by an activated oligonucleotide. Even an unactivated downstream oligonucleotide provides a modest enhancement in the rate of reaction of a primer with a single activated monomer. Here we study the mechanism of these effects through crystallographic studies of RNA complexes with the recently synthesized nonhydrolyzable substrate analog, guanosine 5'-(4-methylimidazolyl)-phosphonate (ICG). ICG mimics 2-methylimidazole activated guanosine-5'-phosphate (2-MeImpG), a commonly used substrate in nonenzymatic primer extension experiments. We present crystal structures of primer-template complexes with either one or two ICG residues bound downstream of a primer. In both cases, the aryl-phosphonate moiety of the ICG adjacent to the primer is disordered. To investigate the effect of a downstream oligonucleotide, we transcribed a short RNA oligonucleotide with either a 5'-ICG residue, a 5'-phosphate or a 5'-hydroxyl. We then determined crystal structures of primer-template complexes with a bound ICG monomer sandwiched between the primer and each of the three downstream oligonucleotides. Surprisingly, all three oligonucleotides rigidify the ICG monomer conformation and position it for attack by the primer 3'-hydroxyl. Furthermore, when GpppG, an analog of the imidazolium-bridged intermediate, is sandwiched between an upstream primer and a downstream helper oligonucleotide, or covalently linked to the 5'-end of the downstream oligonucleotide, the complex is better preorganized for primer extension than in the absence of a downstream oligonucleotide. Our results suggest that a downstream helper oligonucleotide contributes to the catalysis of primer extension by favoring a reactive conformation of the primer-template-intermediate complex.

Dataset Information

Enhanced Nonenzymatic RNA Copying with 2-Aminoimidazole Activated Nucleotides.

Publications

Enhanced Nonenzymatic RNA Copying with 2-Aminoimidazole Activated Nucleotides.

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