Cryomicroscopy reveals nanometer-sized photocopier involved in the origin of life

How the complex molecular machinery of life arose from simple beginnings is a long-standing question. Several lines of evidence point to a primordial “RNA world”, in which an “RNA copying machine” (called a replicase) began making copies of itself and other RNA molecules to restart evolution and life itself. However, the ancient replicase appears to have been lost over time and its role in modern biology has been taken over by more efficient protein machines.

To support the RNA world hypothesis, researchers sought to recreate an equivalent of RNA replicase in the laboratory. Although such molecular “duplicates” of the ancient replicase have been discovered, their detailed molecular structure and mode of action have remained elusive due to the difficulty of determining the structure of dynamic RNA molecules.

Structure of an ice-loving RNA replicase

In a research paper published in Proceedings of the National Academy of Sciencesa team of researchers now reports the first atomic structure of an RNA replicase using cryogenic electron microscopy (cryo-EM).

The RNA replicase studied was developed by the Holliger laboratory (MRC LMB Cambridge, UK) to be efficient in copying long templates using nucleotide triplets in the ice (snow-like) eutectic phase melting).

Returning from postdoctoral studies in the Holliger laboratory, Emil L. Kristoffersen, currently an assistant professor at Aarhus University, facilitated a collaboration with the Andersen laboratory (University of Aarhus, Denmark) to determine the structure of the replicase of RNA by cryo-EM. Interestingly, the structure bears striking similarities to protein-based polymerases, with template-binding, polymerization, and substrate discrimination domains arranged in a molecular shape resembling an open hand.

“It was surprising to find that a ribozyme that we artificially grew in a test tube would exhibit characteristics of natural protein polymerases. This indicates that evolution can discover convergent molecular solutions, regardless of whether they are ‘RNA or protein,’ explains Philipp Holliger, program manager at MRC LMB Cambridge, UK.

Model for RNA synthesis in an RNA world

To better understand how RNA replicase works, the researchers performed a comprehensive mutational study to highlight crucial elements of RNA structure. This analysis confirmed the characteristics of the catalytic site, but also revealed the importance of two so-called kissing loop interactions, which link the scaffold and catalytic subunits, as well as the importance of an RNA domain specific for fidelity, i.e. precision. with which the replicase copies the RNA strands.

Although the researchers were unable to determine the structure of the replicase in action while actively copying RNA, it was possible to construct an RNA-based model of RNA copying that was consistent with all the data experimental.

“Cryo-EM is a powerful method for studying the structure and dynamic characteristics of RNA molecules. By combining cryo-EM data with experiments, we were able to build a model of the inner workings of this complex RNA machine,” explains Ewan McRae, who did cryo-EM work as a postdoc in the Andersen lab at Aarhus University, but has now established his own research group at the Houston Methodist Research Institute in Texas, USA. -United.

Inspiration for RNA nanotechnology and medicine

The study provides an exciting first glimpse of an RNA replicase believed to reside at the very root of the tree of life. Currently developed RNA-based replicases are, however, very inefficient (compared to protein-based polymerases) and cannot yet support their own replication and evolution. The structural insights provided by the reported study could help design more efficient replication mechanisms and thus bring us closer to developing scenarios of the RNA world in the test tube.

“The properties of RNA replicases could be further improved using chemical modifications that could exist in an RNA world. Additionally, research into the origin of life is leading to the discovery of several new building blocks of RNA that could be used in the emerging field of RNA nanotechnology and medicine,” explains Ebbe Sloth Andersen, associate professor at Aarhus University, Denmark.