Structural details of helix-mediated multimerization of the conserved region of TDP-43 C-terminal domain
Azamat Rizuan, Jayakrishna Shenoy, Priyesh Mohanty, Patricia M. dos Passos, José F. Mercado Ortiz, Leanna Bai, Renjith Viswanathan, Julia Zaborowksy, Szu‐Huan Wang, Victoria Johnson, Lohany D. Mamede, Amanda R. Titus, Yuna M. Ayala, Rodolfo Ghirlando, Jeetain Mittal, Nicolas L. Fawzi
Abstract
Pathological inclusions of the C-terminal domain (CTD) of TAR DNA binding protein-43 (TDP-43) are neurodegenerative hallmarks in amyotrophic lateral sclerosis (ALS) and frontotemporal dementia, yet CTD's aggregation propensity complicates structural characterization of native TDP-43. Here we propose structural models for the physiological multimerization of TDP-43 CTD's conserved region (CR) essential for TDP-43 RNA processing. Using NMR spectroscopy, we establish that the native state of TDP-43 CR at physiological conditions is α-helical. Hydrophobic residues drive CR helix-helix assembly, phase separation, and TDP-43 nuclear retention, while polar residues down regulate these processes. An integrative approach combining analytical ultracentrifugation, NMR-derived contacts, AlphaFold2-Multimer modeling, and all-atom molecular dynamics simulations together suggest that TDP-43 CR forms dynamic, multimeric helical assemblies stabilized by a methionine-rich core with specific contributions from a tryptophan/leucine pair. These structures show how ALS-associated mutations disrupt TDP-43 function and provide pharmacologically targetable structures to prevent its conversion into pathogenic β-sheet aggregates.