{"doi":"10.1002/pro.70301","title":"Phosphorylation of <i>Staphylococcus aureus</i> β‐lactamase, a prerequisite to its sequestration on the cytoplasmic membrane","abstract":"Approximately 50% of the class A Staphylococcus aureus β-lactamase (BlaZ) is secreted to the milieu as a resistance vanguard against β-lactam antibiotics. We disclose that the remaining 50% of the β-lactamase is sequestered on the surface of the cytoplasmic membrane in a process that involves its lipidation and phosphorylation, with lipidation being a prerequisite for phosphorylation. Lipidation of a cysteine in the lipobox of BlaZ is ensued by more than one phosphorylation event and cleavage of the signal peptide by a type II signal peptidase. The membrane-bound β-lactamase retains catalytic activity in the turnover of β-lactam antibiotics. A combination of mass-spectrometric analysis and site-directed mutagenesis identified eight sites for phosphorylation. Phosphorylation on Y34, Y41, T151, Y156, T210, and S247 was identified by mass spectrometry, and T119 and S121 by mutagenesis analyses. Individual mutations to alanine variants at Y34, Y41, or T151 prevented BlaZ from phosphorylation altogether, which resulted in the total release of BlaZ into the growth medium. This observation implicates Y34, Y41, and T151 as the primary sites for phosphorylation of BlaZ, which is required for sequestration on the cytoplasmic membrane.","journal":"Protein Science","year":2025,"id":528841,"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":2,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":0.9506,"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":1407128,"name":"Nara Chung","orcid":"0009-0001-9634-3977","position":1,"is_corresponding":false},{"id":1113373,"name":"Hidekazu Hoshino","orcid":"0000-0001-7960-8649","position":2,"is_corresponding":false},{"id":416106,"name":"Matthew M. Champion","orcid":"0000-0001-5927-5277","position":3,"is_corresponding":false},{"id":385035,"name":"Taeok Bae","orcid":"0000-0002-0576-2851","position":4,"is_corresponding":false},{"id":364759,"name":"Shahriar Mobashery","orcid":"0000-0002-7695-7883","position":5,"is_corresponding":false},{"id":739834,"name":"Choon Kim","orcid":"0000-0003-0306-8935","position":0,"is_corresponding":true}],"reference_count":46,"raw_metadata":null,"created_at":"2026-07-19T02:50:52.565868Z","pmid":"40944445","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":[]}