Experimental and Computational Characterization of the Conformational Ensemble and Interaction Motifs of Chiz N-Terminal Intrinsically Disordered Region

Abstract

Cell division of Mycobacterium tuberculosis is crucial for the pathogenesis of tuberculosis. This process is mediated by the divisome, a multi-component, dynamic complex anchored by transmembrane proteins. One such protein is ChiZ, which contains a single transmembrane helix preceded by a 64-residue N-terminal intrinsically disordered region (IDR). We have characterized the conformational ensemble and membrane interactions of the ChiZ IDR by a variety of experimental techniques, including small-angle X-ray scattering (SAXS) and solution and solid-state NMR. 2D 1H-13C INEPT and PRE experiments suggest that the N-terminal IDR of ChiZ remains dynamic in the presence of liposomes, and interactions with the lipid membrane are mediated by arginine residues. To help interpret these data with atomistic details, we have carried out multiple molecular dynamics (MD) simulations in water using several force fields tailored for disordered proteins. The best agreement with SAXS, chemical shift, and NMR relaxation data was obtained by AMBER14SB/Tip4pD, followed closely by AMBER03ws/Tip4p2005s. In the MD simulations, the IDR transiently populated helices, especially in the N-terminal half, including residues 11-15, 19-22, and 28-32. This result explains the lower amide proton exchange rates in the N-terminal half of the IDR observed by solution NMR. Simulations in the presence of a lipid bilayer, in line with the solid-state NMR data, showed multiple arginine residues interacting with the membrane, where any one arginine may engage 2 or 3 lipid headgroups at once. Ongoing work aims to determine the dominant conformations of the full-length ChiZ in the membrane environment.