{"doi":"10.1002/pro.5179","title":"Pathological C‐terminal phosphomimetic substitutions alter the mechanism of liquid–liquid phase separation of TDP‐43 low complexity domain","abstract":"C-terminally phosphorylated TAR DNA-binding protein of 43 kDa (TDP-43) marks the proteinaceous inclusions that characterize a number of age-related neurodegenerative diseases, including amyotrophic lateral sclerosis, frontotemporal lobar degeneration and Alzheimer's disease. TDP-43 phosphorylation at S403/S404 and (especially) at S409/S410 is, in fact, accepted as a biomarker of proteinopathy. These residues are located within the low complexity domain (LCD), which also drives the protein's liquid-liquid phase separation (LLPS). The impact of phosphorylation at these LCD sites on phase separation of the protein is a topic of great interest, as these post-translational modifications and LLPS are both implicated in proteinopathies. Here, we employed a combination of experimental and simulation-based approaches to explore this question on a phosphomimetic model of the TDP-43 LCD. Our turbidity and fluorescence microscopy data show that phosphomimetic Ser-to-Asp substitutions at residues S403, S404, S409 and S410 alter the LLPS behavior of TDP-43 LCD. In particular, unlike the LLPS of unmodified protein, LLPS of the phosphomimetic variants displays a biphasic dependence on salt concentration. Through coarse-grained modeling, we find that this biphasic salt dependence is derived from an altered mechanism of phase separation, in which LLPS-driving short-range intermolecular hydrophobic interactions are modulated by long-range attractive electrostatic interactions. Overall, this in vitro and in silico study provides a physiochemical foundation for understanding the impact of pathologically relevant C-terminal phosphorylation on the LLPS of TDP-43 in a more complex cellular environment.","journal":"Protein Science","year":2024,"id":427920,"datarank":0.0,"base_score":0.0,"endowment":0.0,"self_citation_contribution":0.0,"citation_network_contribution":0.0,"self_endowment_contribution":0.0,"citer_contribution":0.0,"corpus_percentile":null,"corpus_rank":null,"citation_count":14,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":0.9629,"is_data_producer":false,"deposit_databanks":null,"is_oa":true,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":"2024-01-01","fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":1228625,"name":"Brandon Shipley","orcid":"0009-0000-1942-2586","position":1,"is_corresponding":false},{"id":265501,"name":"Krystyna Surewicz","orcid":null,"position":2,"is_corresponding":false},{"id":561633,"name":"Michael Hinczewski","orcid":"0000-0003-2837-7697","position":3,"is_corresponding":false},{"id":235109,"name":"Witold K. Surewicz","orcid":"0000-0001-7940-6027","position":4,"is_corresponding":false},{"id":763960,"name":"Raza Haider","orcid":null,"position":0,"is_corresponding":true}],"reference_count":59,"raw_metadata":null,"created_at":"2026-07-19T01:58:52.576447Z","pmid":"39302099","pmcid":null,"fwci":null,"citation_percentile":null,"influential_citations":0,"oa_status":null,"license":null,"views":0,"total_file_size_bytes":0,"version_count":0,"fair_f":null,"fair_a":null,"fair_i":null,"fair_r":null,"fair_zscore":null,"fair_rationale":null,"fair_model":null,"fair_agent_version":null,"fair_fulltext_source":null,"fair_has_llm":null,"fair_computed_at":null,"clinical_trials":[],"software_tools":[],"db_accessions":[],"linked_datasets":[],"topics":[]}