The structural basis of RNA-catalyzed RNA polymerization

Early life presumably required polymerase ribozymes capable of replicating RNA. Known polymerase ribozymes best approximating such replicases use as their catalytic engine an RNA-ligase ribozyme originally selected from random RNA sequences. Here, we report 3.15 Å crystal structures of this ligase trapped in catalytically viable pre-ligation states, with the 3′-hydroxyl nucleophile positioned for in-line attack on the 5′-triphosphate. Guided by metal and solvent-mediated interactions, the 5′-triphosphate hooks into the major groove of the adjoining RNA duplex in an unanticipated conformation. Two phosphates and the nucleophile jointly coordinate an active-site metal ion. Atomic mutagenesis experiments demonstrate that active-site nucleobase and hydroxyl groups also participate directly in catalysis, collectively playing a role that in proteinaceous polymerases is performed by a second metal ion. Thus artificial ribozymes can employ complex catalytic strategies that differ dramatically from those of analogous biological enzymes.