Abstract
Eukaryotes express at least three nuclear, DNA‐dependent RNA polymerases (Pols I, II, and III) responsible for synthesizing all genome‐encoded RNA. Based on the Pols’ cellular locations, target loci, and transcriptional demands, it is reasonable to hypothesize they have diverged enzymatically over the course of evolution. Since the discovery of the Pols over 50 years ago, the Pols have been largely investigated separately and under disparate experimental conditions, making direct comparisons of the Pols unjustifiable. To directly compare the Pols under identical experimental conditions, we utilized our promoter‐independent in vitro transcription assays. We use a rapid mixing instrument, the chemical quenched‐flow, to analyze single‐ and multi‐nucleotide addition catalyzed by the Pols. We find that Pol I incorporates a single AMP faster than Pols II and III. Interestingly, Pol III incorporates multiple nucleotides faster than Pol I on average. Pol II is consistently the slowest Pol over single‐ and multi‐nucleotide incorporation events. Outside of elongation kinetics, we find additional distinct biochemical properties. Pol II forms the most stable elongation complex of the Pols and is the least error prone Pol. Taken together, our results reveal unique enzymatic characteristics of the Pols that provide new insights into their evolutionary divergence.
Published Version
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call