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Purification and structural characterization of aggregation-prone human TDP-43 involved in neurodegenerative diseases

DOI: 10.1016/j.isci.2020.101159 DOI Help

Authors: Gareth S. A. Wright (University of Liverpool) , Tatiana F. Watanabe (University of Liverpool) , Kangsa Amporndanai (University of Liverpool) , Steven S. Plotkin (University of British Columbia) , Neil R. Cashman (University of British Columbia) , Svetlana V. Antonyuk (University of Liverpool) , S. Samar Hasnain (University of Liverpool)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Iscience

State: Published (Approved)
Published: May 2020

Open Access Open Access

Abstract: Mislocalisation, 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 characterisation has proven 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 NTD domain (residues 1-80) with molecules arranged as parallel spirals with neighbouring molecules arranged in head-to-tail fashion. In order to obtain detergent free, full-length TDP-43 we mutated all six tryptophan residues that provided low-resolution structure of full-length TDP-43 by small angle X-ray scattering and refining the relative positions of individual domains and intrinsically disordered regions in the model using molecular dynamics.

Subject Areas: Biology and Bio-materials

Instruments: B21-High Throughput SAXS , I03-Macromolecular Crystallography

Added On: 20/05/2020 08:43


Discipline Tags:

Neurodegenerative Diseases Non-Communicable Diseases Health & Wellbeing Neurology Structural biology Biophysics Life Sciences & Biotech

Technical Tags:

Diffraction Scattering Macromolecular Crystallography (MX) Small Angle X-ray Scattering (SAXS)