Atomistic Insights into A315E Mutation-Enhanced Pathogenicity of TDP-43 Core Fibrils

Abstract

The aggregation of TAR DNA-binding protein of 43 kDa (TDP-43) into fibrillary deposits is implicated in amyotrophic lateral sclerosis (ALS), and some hereditary mutations localized in the low complexity domain (LCD) facilitate the formation of pathogenic TDP-43 fibrils. A recent cryo-EM study reported the atomic-level structures of the A315E TDP-43 LCD (residues 288–319, TDP-43288–319) core fibril in which the protofilaments have R-shaped structures and hypothesized that A315E U-shaped protofilaments can readily convert to R-shaped protofilaments compared to the wild-type (WT) ones. There are no atomic structures of WT protofilaments available yet. Herein, we performed extensive all-atom explicit-solvent molecular dynamics simulations on A315E and WT protofilaments starting from both the cryo-EM-determined R-shaped and our constructed U-shaped structures. Our simulations show that WT protofilaments also adopt the R-shaped structures but are less stable than their A315E counterparts. Except for R293-E315 salt bridges, N312-F316 hydrophobic interactions and F316–F316 π–π stacking interactions are also crucial for the stabilization of the neck region of the R-shaped A315E protofilaments. The loss of R293-E315 salt bridges and the weakened interactions of N312-F316 and F316–F316 result in the reduced stability of the R-shaped WT protofilaments. Simulations starting from U-shaped folds reveal that A315E protofilaments can spontaneously convert to the cryo-EM-derived R-shaped protofilaments, whereas WT protofilaments convert to R-shape-like structures with remodeled neck regions. The R-shape-like WT protofilaments could act as intermediate states slowing down the U-to-R transition. This study reveals that A315E mutation can not only enhance the structural stability of the R-shaped TDP-43288–319 protofilaments but also promote the U-to-R transition, which provides atomistic insights into the A315E mutation-enhanced TDP-43 pathogenicity in ALS.

ALS-Linked A315T and A315E Mutations Enhance β-Barrel Formation of the TDP-43307–319 Hexamer: A REST2 Simulation Study

“…REST2 can accelerate simulation convergence and enhance the sampling with tremendously reduced computational cost compared with temperature replica exchange molecular dynamics, and it has been widely used in the investigation of the conformational ensembles of intrinsically disordered proteins. − All REST2 simulations in this work were performed using the GROMACS 2018.3 software package patched with the open-source, community-developed PLUMED library, version 2.6.0 . The AMBER99SB-ILDN force field was used in our study in accordance with previous studies showing that this force field can well simulate the aggregation and protofilament stability of fragments of the TDP-43 LCD. ,,,− Each system was placed in a cubic box (8.88 × 8.88 × 8.88 nm 3 ) filled with TIP3P water molecules . Na + and Cl – ions were added in order to neutralize the systems and mimic the physiological salt concentration of 0.15 M. Prior to production REST2 runs, we performed 0.1 ns simulations in the NVT (310 K) ensemble, followed by 0.1 ns simulations in the NPT (310 K, 1 bar) ensemble, to equilibrate the solvent and ions around the protein.…”

“…Lynn et al demonstrated that RRM1/RRM2 exhibit different structural stabilities by performing MD simulations at two different temperatures . By conducting extensive all-atom explicit-solvent MD simulations, we have lately revealed that A315E mutation can not only enhance the structural stability of the R-shaped TDP-43 288–319 protofilaments but also promote the U-to-R transition …”

“…43 By conducting extensive all-atom explicitsolvent MD simulations, we have lately revealed that A315E mutation can not only enhance the structural stability of the Rshaped TDP-43 288−319 protofilaments but also promote the Uto-R transition. 44 Recently, Bowers and co-workers 45 investigated the oligomeric assembly of the TDP-43 307−319 peptide and the effect of A315T/A315E mutation using multiple spectroscopic techniques and reported that A315T and A315E mutants of the TDP-43 307−319 peptide possessed an enhanced capability to form hexamers/octamers, and the hexamers were the toxic agent for these fragments. 45 On the basis of previous findings that β-barrel structures are the toxic oligomeric species of fragments of several proteins, including αB crystallin, 46 amyloid-β, 47 and superoxide dismutase 1 (SOD1), they constructed a β-barrel structure of wild type (WT) and A315T/A315E mutant TDP-43 307−319 by aligning G314 of TDP-43 307−319 with the glycine position (G6) in K11V of αB crystallin (i.e., using the K11V β-barrel of αB crystallin as a template, an out-of-register, antiparallel hexamer) and examined the structural stability of these constructed β-barrels by performing an 80 ns MD simulation.…”

A brief history of amyloid aggregation simulations

“…The structured dimers exhibited different β‐strand arrangements; some of them could be assembled to protofibril models that were found to align with experimental data from H/D exchange NMR spectroscopy and atomic force microscopy. In another study published in 2012, Shea together with Guanghong Wei—who was a postdoctoral researcher in Shea's lab in 2004/2005 and continued to work on amyloid aggregation till today 145–147 —and their coworkers investigated $$$ {\mathrm{A}\upbeta}_{25-35} $$$ oligomers ranging from monomers to tetramers 148 . Their key finding was that the monomer primarily adopted a β‐hairpin conformation, while larger aggregates displayed extended structures.…”

Dissecting the Effect of ALS Mutation G335D on the Early Aggregation of the TDP-43 Amyloidogenic Core Peptide: Helix-to-β-Sheet Transition and Conformational Shift

“…Recently, computational studies are emerging to investigate the conformational properties, aggregation dynamics, and interactions with RNA of fragments of low-complexity domain-containing proteins such as TDP-43 and FUS. ,,− For example, by combining metadynamics and parallel tempering simulations, Fawzi et al found that the TDP-43 310–350 dimer adopts diverse conformations, mainly consisting of parallel/antiparallel helical structures and disordered conformations. By simulating a series of mutant monomers, they found that the G335A/D/N and G338A/D/N mutations can extend the length of the helix .…”