Structural heterogeneity in RNA recognition motif 2 (RRM2) of TAR DNA-binding protein 43 (TDP-43): clue to amyotrophic lateral sclerosis

Abstract

Aberrant misfolding and aggregation of TAR DNA-binding protein 43 (TDP-43) and its fragments have been implicated in amyotrophic lateral sclerosis (ALS) and other neurodegenerative diseases. Within the protein, the second RNA recognition motif (RRM2) has recently been demonstrated to be a major contributor towards aggregation and the resultant toxicity. However, the physicochemical mechanism of its misfolding from the functional folded state is poorly understood. In the present work, we have used a cumulative ∼2µs of molecular dynamics (MD) simulation to study the structural and thermodynamic characteristics of different unfolded intermediates of RRM2 domain of TDP-43. In 6 M GdmCl at 400 K, at RMSD around 1.5 nm, part of the secondary structure i.e. helix still does not melt without significant change in solvent accessibility and intra-protein hydrogen bonds. However, hydrophobic contacts disrupt significantly suggesting that unfolding proceeds through disruption of hydrophobic core of the protein.The temperature dependent free-energy landscapes (FELs) reveal the presence of multiple metastable intermediate states stabilized by hydrophobic (ILV) contacts and hydrogen bonds. These conformational states have all the native helices intact with significant loss of β-sheets. These partially unfolded states are quite compact and characterized by the exposure of aggregation-prone β-sheets, suggesting the increased aggregation propensity of the partially unfolded states. Our results will thus serve to uncover the structural properties of partially unfolded intermediate states that drive TDP-43 misfolding and aggregation. Elucidating the structural characterization of the misfolding and aggregation prone intermediate states of TDP-43 are important to understand its role in ALS and other neurodegenerative diseases.

Communicated by Ramaswamy H. Sarma

Keywords:

• RNA recognition motifs
• protein unfolding
• unfolding intermediates
• hydrophobic contacts
• aggregation

Acknowledgement

Authors sincerely thank DST-SERB for providing the GPU computational Facility.

Author contributions

V.K. designed the study. A.P. performed the experiments and calculations. V.K., A.P. and A.M.L. analyzed the data. V.K. and A.P. wrote the main text with the contributions of A.B., A.M.L and R.P.

Disclosure statement

No potential conflict of interest was reported by the authors.

Additional information

Funding

V.K. and A.P. sincerely thank Science and Engineering Research Board (SERB), Government of India for the award of Young Scientist, SB/YS/LS-161/2014 and YSS/2015/000228/LS.