{"doi":"10.1101/2021.06.14.448324","title":"Reconstitution of the DTX3L-PARP9 complex reveals determinants for high affinity heterodimer formation and enzymatic function","abstract":"Abstract Ubiquitination and ADP-ribosylation are post-translational modifications that play major roles in pathways like DNA damage response and infection, making them attractive targets for therapeutic intervention. DTX3L, an E3 ubiquitin ligase, forms a heterodimer with PARP9. The complex has ubiquitin ligase activity and also ADP-ribosylates the C-terminus of ubiquitin on Gly 76 . NAD + -dependent ADP-ribosylation of ubiquitin by DTX3L-PARP9 prevents ubiquitin from conjugating to protein substrates. By using individually produced proteins, we have studied the interaction between DTX3L and PARP9. We identify that the D3 domain (230 – 510) of DTX3L mediates interaction with PARP9 with nanomolar affinity and an apparent 1:1 stoichiometry. Our results also suggest the formation of a higher molecular weight oligomer mediated by the N-terminus of DTX3L (1-200). Furthermore, we show that ADP-ribosylation of ubiquitin at Gly 76 is a reversible modification that can be removed by several macrodomain-type hydrolases. Our study provides a framework to understand how DTX3L-PARP9 mediates ADP-ribosylation and ubiquitination in an inter-regulatory manner.","journal":"bioRxiv (Cold Spring Harbor Laboratory)","year":2021,"id":221035,"datarank":0.10397207708399181,"base_score":0.6931471805599453,"endowment":0.6931471805599453,"self_citation_contribution":0.10397207708399181,"citation_network_contribution":0.0,"self_endowment_contribution":0.10397207708399181,"citer_contribution":0.0,"corpus_percentile":null,"corpus_rank":null,"citation_count":1,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":0.9569,"is_data_producer":false,"deposit_databanks":null,"is_oa":true,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":"2021-01-01","fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":820771,"name":"Carlos Vela‐Rodríguez","orcid":"0000-0001-6123-1003","position":1,"is_corresponding":false},{"id":681368,"name":"Chunsong Yang","orcid":"0009-0002-9770-5594","position":2,"is_corresponding":false},{"id":820772,"name":"Heli I. Alanen","orcid":"0009-0006-9837-8774","position":3,"is_corresponding":false},{"id":212814,"name":"Fan Liu","orcid":"0000-0002-2358-549X","position":4,"is_corresponding":false},{"id":512007,"name":"Bryce M. Paschal","orcid":"0000-0001-8593-0503","position":5,"is_corresponding":false},{"id":618539,"name":"L. Lehtiö","orcid":"0000-0001-7250-832X","position":6,"is_corresponding":false},{"id":820770,"name":"Yashwanth Ashok","orcid":"0000-0002-5693-6372","position":0,"is_corresponding":true}],"reference_count":55,"raw_metadata":{"citation_network_status":"fetched"},"created_at":"2026-07-18T23:53:50.838581Z","pmid":null,"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":[]}