Purification and Structural Characterization of Aggregation-Prone Human TDP-43 Involved in Neurodegenerative Diseases



Wright, GSA ORCID: 0000-0002-3756-9634, Watanabe, TF, Amporndanai, K, Plotkin, SS, Cashman, NR, Antonyuk, SV ORCID: 0000-0002-2779-9946 and Hasnain, SS
(2020) Purification and Structural Characterization of Aggregation-Prone Human TDP-43 Involved in Neurodegenerative Diseases. iScience, 23 (6).

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Abstract

© 2020 The Author(s) Mislocalization, cleavage, and aggregation of the human protein TDP-43 is found in many neurodegenerative diseases. As is the case with many other proteins that are completely or partially structurally disordered, production of full-length recombinant TDP-43 in the quantities necessary for structural characterization has proved difficult. We show that the full-length TDP-43 protein and two truncated N-terminal constructs 1-270 and 1-263 can be heterologously expressed in E. coli. Full-length TDP-43 could be prevented from aggregation during purification using a detergent. Crystals grown from an N-terminal construct (1-270) revealed only the N-terminal domain (residues 1-80) with molecules arranged as parallel spirals with neighboring molecules arranged in head-to-tail fashion. To obtain detergent-free, full-length TDP-43 we mutated all six tryptophan residues to alanine. This provided sufficient soluble protein to collect small-angle X-ray scattering data. Refining relative positions of individual domains and intrinsically disordered regions against this data yielded a model of full-length TDP-43.

Item Type: Article
Depositing User: Symplectic Admin
Date Deposited: 11 Aug 2020 13:45
Last Modified: 06 Apr 2021 18:10
DOI: 10.1016/j.isci.2020.101159
Open Access URL: https://doi.org/10.1016/j.isci.2020.101159
URI: https://livrepository.liverpool.ac.uk/id/eprint/3097164

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