NMR Fragment-Based Screening against Tandem RNA Recognition Motifs of TDP-43.

Gilbert Nshogoza,Ke Ruan,Yunyu Shi,Rongsheng Ma,Jia Gao,Mingqing Liu,Sayed Ala Moududee,Jiahai Zhang,Fudong Li,Jihui Wu,Yaqian Liu

doi:10.3390/ijms20133230

Abstract

The TDP-43 is originally a nuclear protein but translocates to the cytoplasm in the pathological condition. TDP-43, as an RNA-binding protein, consists of two RNA Recognition Motifs (RRM1 and RRM2). RRMs are known to involve both protein-nucleotide and protein-protein interactions and mediate the formation of stress granules. Thus, they assist the entire TDP-43 protein with participating in neurodegenerative and cancer diseases. Consequently, they are potential therapeutic targets. Protein-observed and ligand-observed nuclear magnetic resonance (NMR) spectroscopy were used to uncover the small molecule inhibitors against the tandem RRM of TDP-43. We identified three hits weakly binding the tandem RRMs using the ligand-observed NMR fragment-based screening. The binding topology of these hits is then depicted by chemical shift perturbations (CSP) of the 15N-labeled tandem RRM and RRM2, respectively, and modeled by the CSP-guided High Ambiguity Driven biomolecular DOCKing (HADDOCK). These hits mainly bind to the RRM2 domain, which suggests the druggability of the RRM2 domain of TDP-43. These hits also facilitate further studies regarding the hit-to-lead evolution against the TDP-43 RRM domain.

Highlights

RNA recognition motifs (RRMs) play diverse roles in post-transcriptional gene expression events such as RNA transport, localization, stability, and mRNA and rRNA processing
The findings demonstrated that transactive response DNA-binding Protein 43kDa (TDP-43) regulates the MALAT1, a non-coding RNA overexpressed in non-small cell lung cancer (NSCLC), through direct binding to MALAT1 RNA at the 3 region by RRM, whose participation is compulsory
TDP-43 tandem RRMs are approximately 160 amino acids long and display a β1α1β2β3α2β4 arrangement of secondary structure, with an additional β-hairpin named β3’β3” [47] or β5 [48,49] which is located between α2β4, and extends the β-sheet surface to be accessible to binding by multiple RNA nucleotides

Summary

Introduction

RNA recognition motifs (RRMs) play diverse roles in post-transcriptional gene expression events such as RNA transport, localization, stability, and mRNA and rRNA processing. RRM is known as the ribonucleoproteins (RNP) domain, as it contains the short and conserved elements RNP1 and RNP2, or RNA binding domain (RBD), that are abundantly distributed in higher vertebrates [1] and ubiquitously found in all kingdoms of life, including viruses and prokaryotes. They participate in important functions such as microRNA biogenesis, apoptosis, and cell division [2,3]. Based on its crucial roles in RNA processing, dysfunctional TDP-43 causes some abnormalities in alternative mRNA splicing, miRNA biogenesis, and RNA-rich granules formation [10]

Methods

Results

Conclusion