{"doi":"10.64898/2025.12.30.697104","title":"The Structural Basis of alpha/beta-tubulin Assembly and Disassembly by Tubulin Cofactors","abstract":"Microtubules polymerize from cytoplasmic pools of soluble αβ-tubulin heterodimers that support diverse cellular functions. The tubulin cofactors, TBCC, TBCD, TBCE, and the Arl2 GTPase, form TBC-DEG assemblies that regulate αβ-tubulin assembly and disassembly from α- and β-tubulins, yet their underlying mechanisms remain incompletely understood. Here, we reconstitute the human TBC-DE and TBC-DEG assemblies from eukaryotic cells co-purified with monomeric β-tubulin intermediates and determine their cryo-EM structures. The structures reveal that TBC-DEG disassembles αβ-tubulin by releasing α-tubulin through a lever-arm-like rotation in TBCE coupled to major conformational change in Arl2 upon its nucleotide release, while TBCD tightly holds β-tubulin. TBCD dissociates α-tubulin by refolding the β-tubulin H10-S8 loop at its intradimer interface. The TBC-DEG-β-tubulin or TBC-DE-β-tubulin assemblies undergo extensive back-to-back dimerization mediated by β-β-tubulin homodimers, formed through their dissociated H8 helices at unoccupied intradimer interfaces. Structural comparisons demonstrate that the TBCE mechanical rotation, driven by the Arl2 GTPase cycle, either delivers α-tubulin or removes it from beneath the TBCD-bound β-tubulin and is directionally regulated by TBCC stabilizing αβ-tubulin interfaces. Our findings suggest that TBC-DEG/TBCC catalyzing heterodimerization of α-tubulin with β-tubulin may have evolved to counteract the β-tubulin intrinsic tendency to form off-pathway toxic homodimers through its exposed α-tubulin-binding intradimer interface.","journal":"bioRxiv (Cold Spring Harbor Laboratory)","year":2025,"id":587513,"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":0.9497,"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":1190613,"name":"Md Ashaduzzaman","orcid":"0000-0002-9967-2530","position":1,"is_corresponding":false},{"id":1503496,"name":"Vishv Gill","orcid":null,"position":2,"is_corresponding":false},{"id":1452404,"name":"Jawdat Al-Bassam","orcid":null,"position":3,"is_corresponding":false},{"id":1326945,"name":"Aryan Taheri","orcid":"0000-0001-8050-8709","position":0,"is_corresponding":true}],"reference_count":28,"raw_metadata":null,"created_at":"2026-07-19T02:59:39.958043Z","pmid":"41509468","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":[]}