{"doi":"10.1101/2023.04.11.536383","title":"Computational design of Matrix Metalloprotenaise-9 (MMP-9) resistant to auto-cleavage","abstract":"Abstract Matrix metalloproteinase-9 (MMP-9) is an endopeptidase that remodels the extracellular matrix and has been implicated as a major driver in cancer metastasis. Hence, there is a high demand for MMP-9 inhibitors for therapeutic purposes. For such drug design efforts, large amounts of MMP-9 are required. Yet, the catalytic domain of MMP-9 (MMP-9 Cat ) is an intrinsically unstable enzyme that tends to auto-cleave within minutes, making it difficult to use in drug design experiments and other biophysical studies. We set our goal to design MMP-9 Cat variant that is active but stable to autocleavage. For this purpose, we first identified potential autocleavage sites on MMP-9 Cat using mass spectroscopy and then eliminated the autocleavage site by predicting mutations that minimize autocleavage potential without reducing enzyme stability. Four computationally designed MMP-9 Cat variants were experimentally constructed and evaluated for auto-cleavage and enzyme activity. Our best variant, Des2, with 2 mutations, was as active as the wild-type enzyme but did not exhibit auto-cleavage after seven days of incubation at 37°C. This MMP-9 Cat variant, with an identical to MMP- 9 Cat WT active site, is an ideal candidate for drug design experiments targeting MMP-9 and enzyme crystallization experiments. The developed strategy for MMP-9 CAT stabilization could be applied to redesign of other proteases to improve their stability for various biotechnological applications.","journal":"bioRxiv (Cold Spring Harbor Laboratory)","year":2023,"id":406320,"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":0.9569,"is_data_producer":false,"deposit_databanks":null,"is_oa":true,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":"2023-01-01","fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":909632,"name":"Solomon Oguche","orcid":null,"position":1,"is_corresponding":false},{"id":714276,"name":"T. Lavy","orcid":"0000-0001-8765-2084","position":2,"is_corresponding":false},{"id":714277,"name":"Oded Kleifeld","orcid":"0000-0003-3091-7154","position":3,"is_corresponding":false},{"id":571574,"name":"Julia M. Shifman","orcid":"0000-0001-6678-6497","position":4,"is_corresponding":false},{"id":756206,"name":"Alessandro Bonadio","orcid":"0000-0001-9003-2732","position":0,"is_corresponding":true}],"reference_count":44,"raw_metadata":null,"created_at":"2026-07-19T01:21:03.432924Z","pmid":"37090502","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":[]}