{"doi":"10.1101/2025.06.02.657148","title":"A histidine switch regulates pH-dependent filament formation by the caspase-9 CARD","abstract":"Abstract The caspase activation and recruitment domain (CARD) mediates protein-protein interactions in apoptotic and inflammatory signaling pathways. In humans, more than 30 proteins contain a CARD, several of which have been reported to polymerize into helical filaments. Here we found that the CARD from the apoptotic protease caspase-9 (C9 CARD ) self assembles into filaments in vitro at physiological pH and salt concentrations. The C9 CARD more readily polymerizes under low-salt or mildly acidic conditions, suggesting a significant role for electrostatic interactions in mediating filament formation. Using NMR spectroscopy, we determined the p K a of the lone histidine residue, H38, which supports a role for histidine protonation in enhancing filament formation. Indeed, mutation of H38 to introduce a positive (H38R) or negative (H38D) charge, or to remove the pH-dependence of the side chain at this site altogether (H38N), dramatically alters the filament propensity of the domain. Using cryo-election microscopy, we determined 3.4- and 3.2-Å structures of the wild-type and H38R C9 CARD filaments, respectively, which provide new insights into the molecular basis of C9 CARD polymerization and its pH dependence via H38.","journal":"bioRxiv (Cold Spring Harbor Laboratory)","year":2025,"id":567520,"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":0,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":0.9596,"is_data_producer":false,"deposit_databanks":null,"is_oa":true,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":"2025-01-01","fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":983173,"name":"Đesika Kolarić","orcid":"0000-0003-3569-6653","position":1,"is_corresponding":false},{"id":475075,"name":"Stefan Bohn","orcid":"0000-0001-9196-622X","position":2,"is_corresponding":false},{"id":1471547,"name":"Dagmar Kolb","orcid":"0000-0002-6387-7446","position":3,"is_corresponding":false},{"id":614120,"name":"Tea Pavkov‐Keller","orcid":"0000-0001-7871-6680","position":4,"is_corresponding":false},{"id":381585,"name":"Iva Pritišanac","orcid":"0000-0002-9152-6555","position":5,"is_corresponding":false},{"id":60679,"name":"Tobias Madl","orcid":"0000-0002-9725-5231","position":6,"is_corresponding":false},{"id":289597,"name":"Ambroise Desfosses","orcid":"0000-0002-6525-5042","position":7,"is_corresponding":false},{"id":381584,"name":"T. Reid Alderson","orcid":"0000-0001-5163-2276","position":8,"is_corresponding":false},{"id":1471875,"name":"Swasti Rawal","orcid":null,"position":0,"is_corresponding":true}],"reference_count":114,"raw_metadata":null,"created_at":"2026-07-19T02:56:44.340853Z","pmid":"40501686","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":[]}