{"doi":"10.1016/j.jbc.2025.110558","title":"Kinetic mechanism of ENPP1 ATPase: Implications for aberrant calcification disorders and enzyme replacement therapy","abstract":"Ectonucleotide Pyrophosphatase Phosphodiesterase 1 (ENPP1) is a transmembrane glycoprotein enzyme with an extracellular catalytic domain that hydrolyzes ATP into AMP and pyrophosphate (PP i ). The ENPP1 ATPase is the major source of extracellular PP i , a critical physiological regulator of calcium phosphate crystal formation and biomineralization. ENPP1 deficiency lowers systemic PP i levels and induces life-threatening arterial calcifications. Enzyme replacement therapy (ERT) with a soluble ENPP1 biologic restores plasma PP i and eliminates calcification and associated mortality in ongoing clinical trials in patients with ENPP1 deficiency. Despite the significant role of ENPP1 in inhibiting vascular calcification and regulating mammalian biomineralization via extracellular PP i levels, little is known about the molecular mechanism of PP i liberation by ENPP1. Here, we provide a kinetic analysis of the ENPP1 catalytic ATPase cycle. Our analysis shows that ATP cleavage, PP i release, and hydrolysis of the covalent AMP-ENPP1 intermediate are rapid (>1000 sec −1 ) and that AMP product release is slow and rate-limiting. The steady-state Michaelis constant of ATP substrate ( K M,T ) is comparable to physiological serum ATP levels of ∼100 nM, rendering ENPP1 activity sensitive to small changes in serum ATP. AMP binds strongly, with an affinity comparable to K M,T , such that ENPP1 is subject to intrinsic regulatory feedback by AMP under physiological concentrations of ∼100 nM. This product inhibition can attenuate ENPP1 during periods of high PP i liberation, maintaining relatively constant plasma PP i levels. The quantitative parameters of the ENPP1 ATPase cycle provided here allow for predictable outcomes of ENPP1 ERT and provide plausible expectations for other PP i -linked calcification disorders.","journal":"Journal of Biological Chemistry","year":2025,"id":526453,"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":3,"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":"2025-01-01","fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":1188937,"name":"Tony Potchernikov","orcid":null,"position":1,"is_corresponding":false},{"id":361092,"name":"Ethan R. Lester","orcid":"0000-0002-7539-0379","position":2,"is_corresponding":false},{"id":361096,"name":"Demetrios T. Braddock","orcid":"0000-0002-0541-4689","position":3,"is_corresponding":false},{"id":308718,"name":"Wenxiang Cao","orcid":"0009-0001-3638-9586","position":4,"is_corresponding":false},{"id":308722,"name":"Enrique M. De La Cruz","orcid":"0000-0003-4798-2892","position":5,"is_corresponding":false},{"id":1402570,"name":"Marisa M Michalchik","orcid":null,"position":0,"is_corresponding":true}],"reference_count":35,"raw_metadata":null,"created_at":"2026-07-19T02:50:30.402772Z","pmid":"40769408","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":[]}