A Clamping Domain Facilitates Processivity and Promoter Recognition in an RNA Polymerase Ribozyme (Razvan COJOCARU)


The “RNA World” proposes that early in evolution RNA could have served both as the carrier of genetic information and as a catalyst. Later in evolution, these RNA functions were gradually replaced by DNA and enzymatic proteins. Arguably, the greatest limitation to exploring this hypothesis is the lack of a true RNA replicase: a processive, trans-acting RNA dependent RNA polymerase ribozyme capable of mediating general replication of RNA. Derived from the Class I ligase ribozyme, we have selected a processive polymerase ribozyme with an RNA clamp domain that mimics many of the mechanisms of modern protein DNA-dependent polymerases.

The selected ribozyme has the following features: 1. The polymerase clamp domain forms an “open” complex when activated with a sigma-like RNA. 2. This “open” complex then searches for a specific RNA promoter. When found, the sigma-like RNA is displaced from the clamp onto the template, triggering a structural rearrangement to a “closed” complex entrained on the RNA template. 3. The displaced sigma-like RNA now serves as a primer, allowing the clamped polymerase to extend a broad range of templates in a processive fashion. This Clamping Polymerase (CP) ribozyme was isolated from a high diversity pool using a range of new in vitro selective strategies. The CP ribozyme has two distinct predicted secondary structures that suggest how the polymerase first binds a sigma-like RNA to find a promoter and then in a second step associates with an RNA template via the formation of a topological clamp.

As an additional feature, this polymerase can synthesize part of its own sigma-like RNA when in the “open” form. This allows part of the polymerase sequence to be encoded into the RNA promoter motif, providing a mechanism early in the evolution of life for a molecular sense of ‘self’ to rapidly evolve. The CP ribozyme demonstrates how the important concepts of promoter recognition, processive polymerization, and self-recognition could have rapidly evolved in a primordial “RNA World”.

Razvan COJOCARU and Peter J. UNRAU, Department of Molecular Biology and Biochemistry, Simon Fraser University, Burnaby, BC, Canada