Synergy among Pausing, Intrinsic Proofreading, and Accessory Proteins Results in Optimal Transcription Speed and...

Cleavage of dinucleotides after the misincorporational pauses serves as a proofreading mechanism that increases transcriptional elongation accuracy. The accuracy is further improved by accessory proteins such as GreA and TFIIS. However, it is not clear why RNAP pauses and why cleavage-factor-assisted proofreading is necessary despite transcriptional errors in vitro being of the same order as those in downstream translation. Here, we developed a chemical-kinetic model that incorporates most relevant features of transcriptional proofreading and uncovers how the balance between speed and accuracy is achieved. We found that long pauses are essential for high accuracy, whereas cleavage-factor-stimulated proofreading optimizes speed. Moreover, in comparison to the cleavage of a single nucleotide or three nucleotides, RNAP backtracking and dinucleotide cleavage improve both speed and accuracy. Our results thereby show how the molecular mechanism and the kinetic parameters of the transcriptional process were evolutionarily optimized to achieve maximal speed and tolerable accuracy.