Insight into the mechanism of nonenzymatic RNA primer extension from the structure of an RNA-GpppG complex

Significance Rudimentary mechanisms of genome replication are essential for the earliest RNA-based cellular life, yet it is unknown how RNA or related polymers could have replicated nonenzymatically. For decades, 2-methylimidazole–activated GMP (2-MeImpG) has been used as a model substrate. We recently showed that two 2-MeImpG monomers react to form an imidazolium-bridged dinucleotide, which then reacts rapidly with the RNA primer. To explore this mechanism, we cocrystallized an RNA primer–template complex with several 5ʹ-5ʹ–linked analogs of the imidazolium-bridged intermediate. The closest analog, GpppG, binds to RNA in a conformation that explains the high reactivity of the imidazolium-bridged intermediate, whereas the structures of other dinucleotide ligands appear less favorable. Our study provides insight into the fundamental mechanism of nonenzymatic RNA self-replication.