Discriminator Effects on Open Complex Formation, Stabilization, and Transcription Initiation

Abstract

The nucleoprotein from the measles virus (MeV) possesses a C-terminal intrinsically disordered domain, referred to as N TAIL , which is essential for virus replication. N TAIL binds to the folded X domain of phosphoprotein P (P XD ), enabling recruitment of the polymerase complex and fine regulation of viral messenger synthesis. N TAIL is highly disordered in the unbound state, and partially folds upon binding to P XD . However, the majority of N TAIL remains disordered in the bound state. The folding upon binding of N TAIL has been characterized using a large number of experimental techniques, revealing many important aspects of binding. However, the role of dynamical reconfiguration of N TAIL disordered regions, and the mechanism by which these contribute to binding remains elusive. To characterize the large-scale dynamical reconfigurations of N TAIL in solution, in the bound and unbound states, we use photo-induced electron transfer (PET) between tryptophan (W) and cysteine (C). This technique allows probing both the W-C distance and the rate of W-C contact formation in IDPs, without using prosthetic dyes. Measured PET rates can be directly compared to results from molecular simulations, providing a more detailed description of IDP conformational ensembles and dynamics. By generating a series of N TAIL W-C mutants, we use PET to probe different regions of the protein, and their modifications upon binding. We compare our experimental results to molecular simulations of N TAIL in the free and bound states. The combined results provide insights into the role of disordered regions, and their conformational dynamics in IDP binding.