{"doi":"10.1021/acschembio.0c00147","title":"Generation of Recombinant Mammalian Selenoproteins\nthrough Genetic Code Expansion with Photocaged Selenocysteine","abstract":"<jats:title>Abstract</jats:title>\n                  <jats:p>Selenoproteins contain the amino acid selenocysteine (Sec) and are found in all domains of life. The functions of many selenoproteins are poorly understood, partly due to difficulties in producing recombinant selenoproteins for cell-biological evaluation. Endogenous mammalian selenoproteins are produced through a noncanonical translation mechanism requiring suppression of the UGA stop codon and a Sec insertion sequence (SECIS) element in the 3′ untranslated region of the mRNA. Here, recombinant selenoproteins are generated in mammalian cells through genetic code expansion, circumventing the requirement for the SECIS element and selenium availability. An engineered orthogonal E. coli leucyl-tRNA synthetase/tRNA pair is used to incorporate a photocaged Sec (DMNB-Sec) at the UAG amber stop codon. DMNB-Sec is successfully incorporated into GFP and uncaged by irradiation of living cells. Furthermore, DMNB-Sec is used to generate the native selenoprotein methionine-R-sulfoxide reductase B1 (MsrB1). Importantly, MsrB1 is shown to be catalytically active after uncaging, constituting the first use of genetic code expansion to generate a functional selenoprotein in mammalian systems. The ability to site-specifically introduce Sec directly in mammalian cells, and temporally modulate selenoprotein activity, will aid in the characterization of mammalian selenoprotein function.</jats:p>","journal":"ACS Chemical\nBiology","year":2020,"id":688851,"datarank":0.519860385419959,"base_score":3.4657359027997265,"endowment":3.4657359027997265,"self_citation_contribution":0.519860385419959,"citation_network_contribution":0.0,"self_endowment_contribution":0.519860385419959,"citer_contribution":0.0,"corpus_percentile":null,"corpus_rank":null,"citation_count":31,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":null,"is_data_producer":false,"deposit_databanks":null,"is_oa":false,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":null,"fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":726120,"name":"Julia A. Falco","orcid":"0000-0002-6890-1476","position":1,"is_corresponding":false},{"id":992119,"name":"Rachel E. Kelemen","orcid":null,"position":2,"is_corresponding":false},{"id":135935,"name":"Masahiro Abo","orcid":null,"position":3,"is_corresponding":false},{"id":752988,"name":"Benjamin V. Chartier","orcid":null,"position":4,"is_corresponding":false},{"id":1799615,"name":"Laura C. Edinger","orcid":null,"position":5,"is_corresponding":false},{"id":460799,"name":"Jingjia Chen","orcid":"0000-0002-6023-1033","position":6,"is_corresponding":false},{"id":117143,"name":"Abhishek Chatterjee","orcid":"0000-0002-6231-5302","position":7,"is_corresponding":false},{"id":135937,"name":"Eranthie Weerapana","orcid":"0000-0002-0835-8301","position":8,"is_corresponding":false},{"id":1799614,"name":"Jennifer\nC. Peeler","orcid":null,"position":0,"is_corresponding":false}],"reference_count":0,"raw_metadata":{"has_enrichment":true,"resolved":true,"title":"Generation of Recombinant Mammalian Selenoproteins\nthrough Genetic Code Expansion with Photocaged Selenocysteine","abstract":"<jats:title>Abstract</jats:title>\n                  <jats:p>Selenoproteins contain the amino acid selenocysteine (Sec) and are found in all domains of life. The functions of many selenoproteins are poorly understood, partly due to difficulties in producing recombinant selenoproteins for cell-biological evaluation. Endogenous mammalian selenoproteins are produced through a noncanonical translation mechanism requiring suppression of the UGA stop codon and a Sec insertion sequence (SECIS) element in the 3′ untranslated region of the mRNA. Here, recombinant selenoproteins are generated in mammalian cells through genetic code expansion, circumventing the requirement for the SECIS element and selenium availability. An engineered orthogonal E. coli leucyl-tRNA synthetase/tRNA pair is used to incorporate a photocaged Sec (DMNB-Sec) at the UAG amber stop codon. DMNB-Sec is successfully incorporated into GFP and uncaged by irradiation of living cells. Furthermore, DMNB-Sec is used to generate the native selenoprotein methionine-R-sulfoxide reductase B1 (MsrB1). Importantly, MsrB1 is shown to be catalytically active after uncaging, constituting the first use of genetic code expansion to generate a functional selenoprotein in mammalian systems. The ability to site-specifically introduce Sec directly in mammalian cells, and temporally modulate selenoprotein activity, will aid in the characterization of mammalian selenoprotein function.</jats:p>","is_dataset_classified":null,"base_score":3.4657359027997265,"endowment":3.4657359027997265,"datacite_reuse_total":0,"file_count":0,"downloads":0,"views":0,"has_version_chain":false,"is_dataset":false,"is_oa":false,"pmid":"32330002","pmcid":"PMC7372505","openalex_id":"https://openalex.org/W3018456911","authors":[],"funders":[{"funder_name":"National Institute of General Medical Sciences","grant_id":"1R35GM134964","title":null},{"funder_name":"National Institute of General Medical Sciences","grant_id":"F32GM131615-01","title":null},{"funder_name":"National Institute of General Medical Sciences","grant_id":"R01GM117004","title":null},{"funder_name":"National Institute of General Medical Sciences","grant_id":"R01GM118431-01A1","title":null},{"funder_name":"National Institute of General Medical Sciences","grant_id":"R01GM124319","title":null},{"funder_name":"National Institute of General Medical Sciences","grant_id":"R01GM126220","title":null},{"funder_name":"NIGMS NIH HHS","grant_id":"R01 GM118431","title":null},{"funder_name":"NIGMS NIH HHS","grant_id":"R35 GM134964","title":null},{"funder_name":"NIGMS NIH HHS","grant_id":"F32 GM131615","title":null},{"funder_name":"National Institutes of Health","grant_id":"5R01GM117004-04","title":"Investigating cysteine PTMs in living cells"},{"funder_name":"National Institutes of Health","grant_id":"5R01GM118431-04","title":"Investigating cysteine-mediated protein activities in C. elegans"},{"funder_name":"National Institutes of Health","grant_id":"1R01GM124319-01","title":"A novel approach to define the roles of oxidative and nitrative post-trasnlational modifications of tryptophan in human biology"},{"funder_name":"National Institutes of Health","grant_id":"1F32GM131615-01","title":"Effect of FABP5 oxidation on fatty acid binding and EGF signaling"},{"funder_name":"National Institutes of Health","grant_id":"1R01GM126220-01A1","title":"A novel strategy to capture post-translational modification-triggered protein-protein interactions"}],"total_grants":14,"fwci":1.9548,"citation_percentile":0.8529169,"influential_citations":0,"citation_trend":[{"year":2019,"count":1},{"year":2020,"count":3},{"year":2021,"count":4},{"year":2022,"count":6},{"year":2023,"count":4},{"year":2024,"count":9},{"year":2025,"count":4}],"oa_status":"green","license":"STM Policy #29","oa_locations":[{"url":"https://www.ncbi.nlm.nih.gov/pmc/articles/7372505","host_type":"repository"},{"url":"https://www.ncbi.nlm.nih.gov/pmc/articles/7372505","host_type":"repository"},{"url":"https://pubs.acs.org/doi/pdf/10.1021/acschembio.0c00147","host_type":"publisher"},{"url":"https://pubs.acs.org/acbcct/article-pdf/15/6/1535/5890199/cb0c00147.pdf","host_type":"publisher"},{"url":"https://doi.org/10.1021/acschembio.0c00147","host_type":"journal"},{"url":"https://pubmed.ncbi.nlm.nih.gov/32330002","host_type":"repository"},{"url":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7372505","host_type":""},{"url":"https://dx.doi.org/10.1021/acschembio.0c00147","host_type":""}],"fields_of_study":["Selenium in Biological Systems","Redox biology and oxidative stress","Trace Elements in Health","0301 basic medicine","03 medical and health sciences","Codon, Terminator","Escherichia coli","Genetic Code","Green Fluorescent Proteins","HEK293 Cells","Humans","Leucine-tRNA Ligase","Protein Engineering","Recombinant Proteins","Selenocysteine","Selenoproteins"],"mesh_terms":["Escherichia coli","Genetic Code","Humans","Leucine-tRNA Ligase","Recombinant Proteins","Protein Engineering","Selenocysteine","Codon, Terminator","Green Fluorescent Proteins","Selenoproteins","HEK293 Cells"],"keywords":["Selenocysteine","Selenoprotein","Genetic code","Stop codon","Transfer RNA","Recombinant DNA","Amino acid","Biology","Biochemistry","Translation (biology)","Aminoacyl tRNA synthetase","Chemistry","Genetics","Cell biology","Gene","Messenger RNA","Enzyme","Glutathione","RNA","Cysteine","HEK293 Cells","Green Fluorescent Proteins","Codon, Terminator","Escherichia coli","Humans","Leucine-tRNA Ligase","Protein Engineering","Selenoproteins","Recombinant Proteins"],"sdg_mappings":[],"linked_datasets":[],"clinical_trials":[],"software_tools":[],"database_accessions":[],"source":"live","citation_network_status":"fetched"},"created_at":"2026-08-19T19:49:27.626971Z","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":[]}