Sequence-dependent correlated segments in the intrinsically disordered region of ChiZ

Abstract

Intrinsically disordered proteins (IDPs) account for a significant fraction of any proteome and are central to numerous cellular functions. Yet how sequences of IDPs code for their conformational dynamics is poorly understood. Here we combined NMR spectroscopy, small-angle X-ray scattering (SAXS), and molecular dynamics (MD) simulations to characterize the conformations and dynamics of ChiZ1-64. This IDP is the N-terminal fragment (residues 1-64) of the transmembrane protein ChiZ, a component of the cell division machinery in Mycobacterium tuberculosis. Its Nhalf contains most of the prolines and all of the anionic residues while the C-half most of the glycines and cationic residues. MD simulations, first validated by SAXS and secondary chemical shift data, found scant a-helices or b-strands but considerable propensity for polyproline II (PPII) torsion angles. Importantly, several blocks of residues (e.g., 11-29) emerge as "correlated segments", identified by frequent formation of PPII stretches, salt bridges, cation-p interactions, and sidechain-backbone hydrogen bonds. NMR relaxation experiments showed non-uniform transverse relaxation rates (R2s) and nuclear Overhauser enhancements (NOEs) along the sequence (e.g., high R2s and NOEs for residues 11-14 and 23-28). MD simulations further revealed that the extent of segmental correlation is sequence-dependent: segments where internal interactions are more prevalent manifest elevated "collective" motions on the 5-10 ns timescale and suppressed local motions on the sub-ns timescale. Amide proton exchange rates provides corroboration, with residues in the most correlated segment exhibiting the highest protection factors. We propose correlated segment as a defining feature for the conformation and dynamics of IDPs.

Fuzzy Association of an Intrinsically Disordered Protein with Acidic Membranes

“…In full-length ChiZ (ChiZ-FL), NT is followed by a 21-residue transmembrane helix; on the periplasmic side, a C-terminal LysM domain (residues 113–165) is connected to the transmembrane helix by a 26-residue linker ( Figure 1 ). In a previous study, 20 we showed that, in solution, the NT-only construct ChiZ1-64 is fully disordered without detectable α-helix or β-sheet formation, but with polyproline II (PPII) formation and intramolecular interactions including salt bridges concentrated in the first half of the sequence. Here, we investigated NT-membrane association by solution NMR in the context of ChiZ1-64 and by solid-state NMR on both ChiZ1-64 and ChiZ-FL.…”

Extreme Fuzzy Association of an Intrinsically Disordered Protein with Acidic Membranes

“…1). In a previous study 20 , we showed that, in solution, the NT-only construct, ChiZ1-64, is fully disordered without detectable α-helix or b-sheet formation, but with polyproline II (PPII) formation and intramolecular interactions including salt bridges concentrated in the first half of the sequence. Here we investigated NT-membrane association by solution NMR in the context of ChiZ1-64 and by solidstate NMR on both ChiZ1-64 and ChiZ-FL.…”

“…Association is indicated by loss of crosspeaks due to line broadening from the slow tumbling when a residue interacts with a liposome. Relative to the 1 H-15 N HSQC spectrum of ChiZ1-64 in solution (hereafter "unbound" ChiZ1-64) 20 , no major loss in NMR signals was detected when the liposomes contained POPC only ( Fig. 2a), 4:1 DOPC:DOPE (Fig.…”

“…Both in unbound ChiZ1-64 20 and in the INEPT spectrum of POPG:POPE-bound ChiZ1-64, the Arg Ca-Cb pair has two distinct crosspeaks, one at (56.3, 30.8) (chemical shifts in ppm), and the other at (54.0, 30.4) ( Fig. 3b, black contours).…”

“…The two halves of unbound ChiZ1-64 are asymmetric not only in salt-bridge formation but also in PPII propensity (there are very little propensities for helices and bstrands; Supplementary Fig. 9) 20 . Three PPII stretches form with high probabilities (>50%), all in N-half: V4RP6, P10DP12, and A27EP29.…”