Abstract
Non-enzymatic RNA self-replication is integral to the emergence of the 'RNA World'. Despite considerable progress in non-enzymatic template copying, demonstrating a full replication cycle remains challenging due to the difficulty of separating the strands of the product duplex. Here, we report a prebiotically plausible approach to strand displacement synthesis in which short 'invader' oligonucleotides unwind an RNA duplex through a toehold/branch migration mechanism, allowing non-enzymatic primer extension on a template that was previously occupied by its complementary strand. Kinetic studies of single-step reactions suggest that following invader binding, branch migration results in a 2:3 partition of the template between open and closed states. Finally, we demonstrate continued primer extension with strand displacement by employing activated 3'-aminonucleotides, a more reactive proxy for ribonucleotides. Our study suggests that complete cycles of non-enzymatic replication of the primordial genetic material may have been facilitated by short RNA oligonucleotides.
Highlights
The replication of a genetic polymer within a vesicle capable of growth and division should lead to a protocell able to undergo Darwinian evolution
This experiment demonstrates that primer extension by strand displacement synthesis can be catalyzed by short RNA oligonucleotides
Our experiments show that non-enzymatic primer extension by strand displacement synthesis can be catalyzed by short RNA oligonucleotides, resulting in the template-directed synthesis of RNA
Summary
The replication of a genetic polymer within a vesicle capable of growth and division should lead to a protocell able to undergo Darwinian evolution. The construction of a protocell capable of autonomous reproduction, even within an artificial laboratory setting, may lead to significant insights into the origin of life (Joyce & Szostak, 2018). Prior to the emergence of enzymes or ribozymes on the early Earth, the first genetic material had to rely on non-enzymatic copying reactions to accomplish self-replication. Following template-directed RNA copying, the daughter strand, which is complementary to the template, must undergo another round of copying to generate a product with the same sequence as the original template, completing the replication process. Recent discoveries have greatly improved the rate and extent of chemical RNA copying reactions in laboratory studies (Li et al, 2017; O’Flaherty et al, 2018). After the first round of templated copying, the daughter strand is sequestered within a stable duplex with its parent strand
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