{"doi":"10.1101/2021.09.22.461393","title":"Cell-surface tethered promiscuous biotinylators enable small-scale surface proteomics of human exosomes","abstract":"Abstract Characterization of cell surface proteome differences between cancer and healthy cells is a valuable approach for the identification of novel diagnostic and therapeutic targets. However, selective sampling of surface proteins for proteomics requires large samples (>10e7 cells) and long labeling times. These limitations preclude analysis of material-limited biological samples or the capture of rapid surface proteomic changes. Here, we present two labeling approaches to tether exogenous peroxidases (APEX2 and HRP) directly to cells, enabling rapid, small-scale cell surface biotinylation without the need to engineer cells. We used a novel lipidated DNA-tethered APEX2 (DNA-APEX2), which upon addition to cells promoted cell agnostic membrane-proximal labeling. Alternatively, we employed horseradish peroxidase (HRP) fused to the glycan binding domain of wheat germ agglutinin (WGA-HRP). This approach yielded a rapid and commercially inexpensive means to directly label cells containing common N-Acetylglucosamine (GlcNAc) and sialic acid glycans on their surface. The facile WGA-HRP method permitted high surface coverage of cellular samples and enabled the first comparative surface proteome characterization of cells and cell-derived exosomes, leading to the robust quantification of 1,020 cell and exosome surface proteins. We identified a newly-recognized subset of exosome-enriched markers, as well as proteins that are uniquely upregulated on Myc oncogene-transformed prostate cancer exosomes. These two cell-tethered enzyme surface biotinylation approaches are highly advantageous for rapidly and directly labeling surface proteins across a range of material-limited sample types.","journal":"bioRxiv (Cold Spring Harbor Laboratory)","year":2021,"id":221681,"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":1,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":0.9462,"is_data_producer":false,"deposit_databanks":null,"is_oa":true,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":"2021-01-01","fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":303659,"name":"Susanna K. Elledge","orcid":"0000-0002-9621-3881","position":1,"is_corresponding":false},{"id":821807,"name":"Jiuling Yang","orcid":"0000-0002-5301-6976","position":2,"is_corresponding":false},{"id":274317,"name":"James R. Byrnes","orcid":"0000-0003-0297-1209","position":3,"is_corresponding":false},{"id":237508,"name":"Jeff E. Glasgow","orcid":"0000-0001-6699-8983","position":4,"is_corresponding":false},{"id":821808,"name":"Robert Blelloch","orcid":"0000-0002-1975-0798","position":5,"is_corresponding":false},{"id":237520,"name":"James A. Wells","orcid":"0000-0001-8267-5519","position":6,"is_corresponding":false},{"id":292905,"name":"Lisa L. Kirkemo","orcid":"0000-0003-1686-6987","position":0,"is_corresponding":true}],"reference_count":77,"raw_metadata":null,"created_at":"2026-07-18T23:53:55.215284Z","pmid":null,"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":[]}