{"doi":"10.1101/2025.10.21.683677","title":"The nucleotide exchange factor, GrpE, modulates substrate affinity by interaction of its N-terminal tails with the DnaK substrate-binding domain","abstract":"Abstract The 70-kDa heat shock proteins (Hsp70s) assist in protein folding through allosteric communication between their nucleotide-binding domains (NBDs) and substrate-binding domains (SBDs), which are connected by an interdomain linker. Their nucleotide-dependent allosteric cycle is modulated by ligand binding and co-chaperones, including nucleotide exchange factors (NEFs). GrpE, the NEF for the E. coli Hsp70, DnaK, has been proposed to have a dual effect on the chaperone, facilitating the exchange of ADP for ATP in the NBD in a temperature-dependent fashion and promoting substrate release from the SBD. We recently reported NMR-based evidence that GrpE binding to DnaK has a direct structural effect on the SBD. Here, we expanded on these findings and obtained new evidence for a model in which the disordered N-terminal tails of GrpE facilitate peptide dissociation from the nucleotide-free DnaK/GrpE complex by transiently binding to the canonical substrate-binding site in the SBD. This GrpE/SBD interaction, while weak, is favored by the high local concentration of the tails around the SBD after complex formation. Moreover, we identified the DnaK binding motif in GrpE’s N-terminal disordered tails as 17 IIM 19 , which is conserved across many bacterial species. Excitingly, our data further suggest a mechanism for the temperature-dependence of GrpE’s modulation of DnaK’s refolding activity: as the temperature increases, unfolding of GrpE’s coiled-coil weakens its contacts with the SBD, reducing N-terminal tail binding, and thus increasing DnaK affinity to substrates.","journal":"bioRxiv (Cold Spring Harbor Laboratory)","year":2025,"id":578724,"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":0.9547,"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":306837,"name":"María-Agustina Rossi","orcid":"0000-0003-4720-4070","position":1,"is_corresponding":false},{"id":734645,"name":"Eugenia M. Clérico","orcid":"0000-0002-4433-8842","position":2,"is_corresponding":false},{"id":410047,"name":"Robert V. Williams","orcid":"0000-0003-1839-6122","position":3,"is_corresponding":false},{"id":612912,"name":"Lila M. Gierasch","orcid":"0000-0002-5706-115X","position":4,"is_corresponding":false},{"id":1488640,"name":"Akshitha Maqtedar","orcid":"0009-0004-3811-1475","position":0,"is_corresponding":true}],"reference_count":36,"raw_metadata":{"citation_network_status":"fetched"},"created_at":"2026-07-19T02:58:24.957414Z","pmid":"41279868","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":[]}