Proline isomerization compensates for the effect of phosphorylation in conformations of the disordered RNA polymerase II CTD.

Abstract

RNA Polymerase II transcribes mRNA. The largest subunit of this complex contains a C-terminal domain (CTD) that is an intrinsically disordered protein with a repetitive amino acid heptad sequence making the domain very difficult to study. Kinases and Phosphatases regulate CTD through post-translational modifications. Small angle x-ray scattering data shows very little change in CTD compaction with phosphorylation, despite the repulsion between the negatively charged phosphate groups. Data also shows an increase in Pro6 isomerization when Ser5 is phosphorylated. Prior work shows that a variation of the consensus sequence where Ser7 is replaced with Asn increases the population of the proline cis conformation. We use Gaussian accelerated molecular dynamics simulations to determine how the CTD domain conformation and dynamics are impacted by phosphorylation. Simulations were run on a 21-residue construct consisting of three heptads, with the middle heptad containing Asn7. Results show that due to proline isomerization, the domain can move reversibly between a compact and extended structure. The phosphorylated CTD domain samples more prolines in cis, which compensates for the effect of phosphorylation, resulting in a similar radius of gyration to the unphosphorylated domain. Future work includes running simulations with the consensus sequence for all three heptads of our construct to determine why having asparagine in position 7 increases Pro6 isomerization compared to serine.