{"doi":"10.1016/j.xpro.2024.103483","title":"Protocol for purifying and reconstituting a vacuole membrane transporter Ypq1 into proteoliposomes","abstract":"Studying the biochemical function of membrane transporters is important in understanding the biology of transporter-laden organelles such as lysosomes and vacuoles. We present a protocol for overexpressing, purifying, and reconstituting a vacuole membrane transporter Ypq1 into proteoliposomes and describe steps to measure transport activity using radioactive substrates. The protocols established here can be used to study other vacuolar or lysosomal membrane transporters. For complete details on the use and execution of this protocol, please refer to Arines et al. 1 • Steps for affinity chromatography and FPLC for protein purification • Instructions for characterizing activity with radioactive substrate • Guidance on reconstituting protein into a liposome system Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics. Studying the biochemical function of membrane transporters is important in understanding the biology of transporter-laden organelles such as lysosomes and vacuoles. We present a protocol for overexpressing, purifying, and reconstituting a vacuole membrane transporter Ypq1 into proteoliposomes and describe steps to measure transport activity using radioactive substrates. The protocols established here can be used to study other vacuolar or lysosomal membrane transporters.","journal":"STAR Protocols","year":2024,"id":474741,"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":0.958,"is_data_producer":false,"deposit_databanks":null,"is_oa":true,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":"2024-01-01","fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":1154190,"name":"Aleksander Wielenga","orcid":null,"position":1,"is_corresponding":false},{"id":347897,"name":"Randy B Stockbridge","orcid":"0000-0001-8848-3032","position":2,"is_corresponding":false},{"id":319836,"name":"Ming Li","orcid":"0000-0002-1247-2377","position":3,"is_corresponding":false},{"id":319832,"name":"Felichi Mae Arines","orcid":"0000-0002-5770-3116","position":0,"is_corresponding":true}],"reference_count":17,"raw_metadata":{"citation_network_status":"fetched"},"created_at":"2026-07-19T02:06:13.042906Z","pmid":"39661504","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":[]}