Millisecond Functional Dynamics of RNA Polymerases Elucidated by Markov State Models

RNA polymerase II (Pol II) undergoes a series of conformational changes when actively transcribing genes. I will present our group’s work on constructing Markov State Models (MSMs) from molecular dynamics (MD) simulations to elucidate dynamics of these conformational changes, including forward translocation, backtracking, and pyrophosphate release. In particular, I will explain the molecular mechanisms for Pol II to maintain transcription fidelity. Pol II efficiently detects errors and backtracks. Using MSMs, we identified a critical threonine residue acting as the checkpoint for backtracking. Following backtracking, Pol II cleaves the backtracked mis-incorporated nucleotide. We found strikingly that while specific amino acid residues of Pol II are critical for backtracking, cleavage of the mis-incorporated nucleotide only requires the RNA nucleotide itself (i.e., phosphate oxygen of mis-incorporated nucleotide). Based combining computational and experimental work, we reveal how Pol II accomplishes the task to catalyze two distinct chemical reactions using a single active site in a coordinated fashion, which is longstanding question in transcription. In addition, I will present our recent work on developing Generalized Master Equation (GME) models that encodes the non-Markovian dynamics in a generally time-dependent memory kernel. We successfully applied GMEs to elucidate molecular mechanisms of DNA loading into a bacterial RNA polymerase complex via flexible loading gate (consisting of the clamp and β-lobe domain), a process occurs at millisecond. We further reveal the mechanism of an antibiotics targeting this bacterial RNA Polymerase.