{"doi":"10.1093/mtomcs/mfac014","title":"Characterization of the nickel-inserting cyclometallase LarC from <i>Moorella thermoacetica</i> and identification of a cytidinylylated reaction intermediate","abstract":"LarC catalyzes the CTP-dependent insertion of nickel ion into pyridinium-3,5-bisthiocarboxylic acid mononucleotide (P2TMN), the final biosynthetic step for generating the nickel-pincer nucleotide (NPN) enzyme cofactor. In this study, we characterized a LarC homolog from Moorella thermoacetica (LarCMt) and characterized selected properties of the protein. We ruled out the hypothesis that enzyme inhibition by its product pyrophosphate accounts for its apparent single-turnover activity. Most notably, we identified a cytidinylylated-substrate intermediate that is formed during the reaction of LarCMt. Selected LarCMt variants with substitutions at the predicted CTP-binding site retained substantial amounts of activity, but exhibited greatly reduced levels of the CMP-P2TMN intermediate. In contrast, enhanced amounts of the CMP-P2TMN intermediate were generated when using LarCMt from cells grown on medium without supplemental nickel. On the basis of these results, we propose a functional role for CTP in the unprecedented nickel-insertase reaction during NPN biosynthesis.","journal":"Metallomics","year":2022,"id":268835,"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":18,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":0.9573,"is_data_producer":false,"deposit_databanks":null,"is_oa":true,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":"2022-01-01","fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":334698,"name":"Jian Hu","orcid":"0000-0001-6657-9826","position":1,"is_corresponding":false},{"id":475596,"name":"Robert P. Hausinger","orcid":"0000-0002-3643-2054","position":2,"is_corresponding":false},{"id":475595,"name":"Aiko Turmo","orcid":"0000-0002-6380-8811","position":0,"is_corresponding":true}],"reference_count":18,"raw_metadata":null,"created_at":"2026-07-19T00:27:14.088663Z","pmid":"35225337","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":[]}