{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"bioRxiv"}],"indexed":{"date-parts":[[2026,1,15]],"date-time":"2026-01-15T20:11:14Z","timestamp":1768507874810,"version":"3.49.0"},"posted":{"date-parts":[[2023,9,18]]},"group-title":"Biochemistry","reference-count":79,"publisher":"openRxiv","content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2023,9,18]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                <jats:p>\n                  Encapsulins are self-assembling protein nanocompartments capable of selectively encapsulating dedicated cargo proteins, including enzymes involved in iron storage, sulfur metabolism, and stress resistance. They represent a unique compartmentalization strategy used by many pathogens to facilitate specialized metabolic capabilities. Encapsulation is mediated by specific cargo protein motifs known as targeting peptides (TPs), though the structural basis for encapsulation of the largest encapsulin cargo class, dye-decolorizing peroxidases (DyPs), is currently unknown. Here, we characterize a DyP-containing encapsulin from the enterobacterial pathogen\n                  <jats:italic>Klebsiella pneumoniae<\/jats:italic>\n                  . By combining cryo-electron microscopy with TP mutagenesis, we elucidate the molecular basis for cargo encapsulation. TP binding is mediated by cooperative hydrophobic and ionic interactions as well as shape complementarity. Our results expand the molecular understanding of enzyme encapsulation inside protein nanocompartments and lay the foundation for rationally modulating encapsulin cargo loading for biomedical and biotechnological applications.\n                <\/jats:p>","DOI":"10.1101\/2023.09.18.558302","type":"posted-content","created":{"date-parts":[[2023,9,19]],"date-time":"2023-09-19T00:05:16Z","timestamp":1695081916000},"source":"Crossref","is-referenced-by-count":2,"title":["Structural basis for peroxidase encapsulation in a protein nanocompartment"],"prefix":"10.64898","author":[{"ORCID":"https:\/\/orcid.org\/0000-0001-8006-5661","authenticated-orcid":false,"given":"Jesse A.","family":"Jones","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8871-3053","authenticated-orcid":false,"given":"Michael P.","family":"Andreas","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-6328-2031","authenticated-orcid":false,"given":"Tobias W.","family":"Giessen","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"54368","reference":[{"key":"2023092204350854000_2023.09.18.558302v1.1","doi-asserted-by":"publisher","DOI":"10.1016\/j.ceb.2013.12.007"},{"key":"2023092204350854000_2023.09.18.558302v1.2","doi-asserted-by":"publisher","DOI":"10.1042\/BJ20120957"},{"key":"2023092204350854000_2023.09.18.558302v1.3","doi-asserted-by":"publisher","DOI":"10.1016\/j.biochi.2015.03.021"},{"key":"2023092204350854000_2023.09.18.558302v1.4","doi-asserted-by":"publisher","DOI":"10.1021\/jacs.5b11734"},{"key":"2023092204350854000_2023.09.18.558302v1.5","doi-asserted-by":"publisher","DOI":"10.1146\/annurev.micro.112408.134211"},{"key":"2023092204350854000_2023.09.18.558302v1.6","doi-asserted-by":"crossref","first-page":"36","DOI":"10.1016\/j.mib.2021.05.010","article-title":"Bacterial microcompartments: tiny organelles with big potential","volume":"63","year":"2021","journal-title":"Curr Opin Microbiol"},{"key":"2023092204350854000_2023.09.18.558302v1.7","doi-asserted-by":"crossref","unstructured":"Giessen, T.W . Encapsulins. Annu Rev Biochem (2022).","DOI":"10.1146\/annurev-biochem-040320-102858"},{"key":"2023092204350854000_2023.09.18.558302v1.8","doi-asserted-by":"publisher","DOI":"10.1016\/j.mib.2021.05.009"},{"key":"2023092204350854000_2023.09.18.558302v1.9","doi-asserted-by":"publisher","DOI":"10.1016\/j.pbi.2016.03.009"},{"key":"2023092204350854000_2023.09.18.558302v1.10","doi-asserted-by":"crossref","first-page":"219","DOI":"10.1111\/1462-2920.16283","article-title":"Role of carboxysomes in cyanobacterial CO(2) assimilation: CO(2) concentrating mechanisms and metabolon implications","volume":"25","year":"2023","journal-title":"Environ Microbiol"},{"key":"2023092204350854000_2023.09.18.558302v1.11","doi-asserted-by":"crossref","unstructured":"Turmo, A. , Gonzalez-Esquer, C.R. & Kerfeld, C.A . Carboxysomes: metabolic modules for CO2 fixation. FEMS Microbiol Lett 364(2017).","DOI":"10.1093\/femsle\/fnx176"},{"key":"2023092204350854000_2023.09.18.558302v1.12","doi-asserted-by":"publisher","DOI":"10.1016\/j.mib.2021.05.011"},{"key":"2023092204350854000_2023.09.18.558302v1.13","doi-asserted-by":"crossref","first-page":"28","DOI":"10.1016\/j.mib.2021.04.008","article-title":"Ethanolamine bacterial microcompartments: from structure, function studies to bioengineering applications","volume":"62","year":"2021","journal-title":"Curr Opin Microbiol"},{"key":"2023092204350854000_2023.09.18.558302v1.14","doi-asserted-by":"publisher","DOI":"10.1007\/s00253-005-0295-0"},{"key":"2023092204350854000_2023.09.18.558302v1.15","doi-asserted-by":"crossref","first-page":"4377","DOI":"10.1039\/D3TB00288H","article-title":"Encapsulin cargo loading: progress and potential","volume":"11","year":"2023","journal-title":"J Mater Chem B"},{"key":"2023092204350854000_2023.09.18.558302v1.16","doi-asserted-by":"crossref","first-page":"35","DOI":"10.1134\/S0006297923010042","article-title":"Encapsulins: Structure, Properties, and Biotechnological Applications","volume":"88","year":"2023","journal-title":"Biochemistry (Mosc)"},{"key":"2023092204350854000_2023.09.18.558302v1.17","doi-asserted-by":"crossref","unstructured":"Giessen, T.W. et al. Large protein organelles form a new iron sequestration system with high storage capacity. Elife 8(2019).","DOI":"10.7554\/eLife.46070"},{"key":"2023092204350854000_2023.09.18.558302v1.18","doi-asserted-by":"crossref","first-page":"551","DOI":"10.1016\/j.str.2022.01.008","article-title":"Structural characterization of the Myxococcus xanthus encapsulin and ferritin-like cargo system gives insight into its iron storage mechanism","volume":"30","year":"2022","journal-title":"Structure"},{"key":"2023092204350854000_2023.09.18.558302v1.19","doi-asserted-by":"crossref","unstructured":"Nichols, R.J. et al. Discovery and characterization of a novel family of prokaryotic nanocompartments involved in sulfur metabolism. Elife 10(2021).","DOI":"10.7554\/eLife.59288"},{"key":"2023092204350854000_2023.09.18.558302v1.20","doi-asserted-by":"crossref","unstructured":"Benisch, R. , Andreas, M.P. & Giessen, T.W. A widespread proteinaceous sulfur storage compartment in bacteria. bioRxiv (2023).","DOI":"10.1101\/2023.06.21.545984"},{"key":"2023092204350854000_2023.09.18.558302v1.21","doi-asserted-by":"crossref","unstructured":"Tang, Y. et al. Cryo-EM structure of Mycobacterium smegmatis DyP-loaded encapsulin. Proc Natl Acad Sci U S A 118(2021).","DOI":"10.1073\/pnas.2025658118"},{"key":"2023092204350854000_2023.09.18.558302v1.22","doi-asserted-by":"publisher","DOI":"10.1074\/jbc.M114.570119"},{"key":"2023092204350854000_2023.09.18.558302v1.23","doi-asserted-by":"publisher","DOI":"10.1080\/10409238.2017.1337709"},{"key":"2023092204350854000_2023.09.18.558302v1.24","doi-asserted-by":"publisher","DOI":"10.1038\/nsmb.1473"},{"key":"2023092204350854000_2023.09.18.558302v1.25","doi-asserted-by":"publisher","DOI":"10.1016\/j.coviro.2019.02.001"},{"key":"2023092204350854000_2023.09.18.558302v1.26","doi-asserted-by":"crossref","first-page":"4748","DOI":"10.1038\/s41467-021-25071-y","article-title":"Large-scale computational discovery and analysis of virus-derived microbial nanocompartments","volume":"12","year":"2021","journal-title":"Nat Commun"},{"key":"2023092204350854000_2023.09.18.558302v1.27","doi-asserted-by":"publisher","DOI":"10.1038\/s41598-019-56533-5"},{"key":"2023092204350854000_2023.09.18.558302v1.28","doi-asserted-by":"crossref","first-page":"101787","DOI":"10.1016\/j.mex.2022.101787","article-title":"Heterologous expression and purification of encapsulins in Streptomyces coelicolor","volume":"9","year":"2022","journal-title":"MethodsX"},{"key":"2023092204350854000_2023.09.18.558302v1.29","doi-asserted-by":"publisher","DOI":"10.1038\/s41598-021-84329-z"},{"key":"2023092204350854000_2023.09.18.558302v1.30","doi-asserted-by":"crossref","unstructured":"Gabashvili, A.N. et al. Myxococcus xanthus Encapsulin as a Promising Platform for Intracellular Protein Delivery. Int J Mol Sci 23(2022).","DOI":"10.3390\/ijms232415591"},{"key":"2023092204350854000_2023.09.18.558302v1.31","doi-asserted-by":"crossref","first-page":"231","DOI":"10.1016\/j.synbio.2021.09.001","article-title":"Bioengineering bacterial encapsulin nanocompartments as targeted drug delivery system","volume":"6","year":"2021","journal-title":"Synth Syst Biotechnol"},{"key":"2023092204350854000_2023.09.18.558302v1.32","doi-asserted-by":"publisher","DOI":"10.1016\/j.vaccine.2018.05.034"},{"key":"2023092204350854000_2023.09.18.558302v1.33","doi-asserted-by":"publisher","DOI":"10.1002\/bit.25159"},{"key":"2023092204350854000_2023.09.18.558302v1.34","doi-asserted-by":"crossref","first-page":"857","DOI":"10.1021\/acssynbio.0c00636","article-title":"Encapsulin Nanocontainers as Versatile Scaffolds for the Development of Artificial Metabolons","volume":"10","year":"2021","journal-title":"ACS Synth Biol"},{"key":"2023092204350854000_2023.09.18.558302v1.35","doi-asserted-by":"crossref","first-page":"3504","DOI":"10.1021\/acssynbio.2c00391","article-title":"Engineered Protein Nanocages for Concurrent RNA and Protein Packaging In Vivo","volume":"11","year":"2022","journal-title":"ACS Synth Biol"},{"key":"2023092204350854000_2023.09.18.558302v1.36","doi-asserted-by":"crossref","first-page":"e1685","DOI":"10.1002\/wnan.1685","article-title":"Engineering spatiotemporal organization and dynamics in synthetic cells","volume":"13","year":"2021","journal-title":"Wiley Interdiscip Rev Nanomed Nanobiotechnol"},{"key":"2023092204350854000_2023.09.18.558302v1.37","doi-asserted-by":"crossref","first-page":"17029","DOI":"10.1038\/nmicrobiol.2017.29","article-title":"Widespread distribution of encapsulin nanocompartments reveals functional diversity","volume":"2","year":"2017","journal-title":"Nat Microbiol"},{"key":"2023092204350854000_2023.09.18.558302v1.38","doi-asserted-by":"crossref","unstructured":"Lien, K.A. et al. A nanocompartment system contributes to defense against oxidative stress in Mycobacterium tuberculosis. Elife 10(2021).","DOI":"10.7554\/eLife.74358"},{"key":"2023092204350854000_2023.09.18.558302v1.39","doi-asserted-by":"crossref","first-page":"751","DOI":"10.1002\/bab.2015","article-title":"Biochemical features of dye-decolorizing peroxidases: Current impact on lignin degradation","volume":"67","year":"2020","journal-title":"Biotechnol Appl Biochem"},{"key":"2023092204350854000_2023.09.18.558302v1.40","doi-asserted-by":"crossref","unstructured":"Sugano, Y. & Yoshida, T . DyP-Type Peroxidases: Recent Advances and Perspectives. Int J Mol Sci 22(2021).","DOI":"10.3390\/ijms22115556"},{"key":"2023092204350854000_2023.09.18.558302v1.41","doi-asserted-by":"crossref","first-page":"38","DOI":"10.32607\/20758251-2011-3-4-38-54","article-title":"NAD (+) -dependent Formate Dehydrogenase from Plants","volume":"3","year":"2011","journal-title":"Acta Naturae"},{"key":"2023092204350854000_2023.09.18.558302v1.42","doi-asserted-by":"crossref","first-page":"544","DOI":"10.1107\/S2052252523006437","article-title":"Structure and function relationship of formate dehydrogenases: an overview of recent progress","volume":"10","year":"2023","journal-title":"IUCrJ"},{"key":"2023092204350854000_2023.09.18.558302v1.43","doi-asserted-by":"publisher","DOI":"10.1016\/j.copbio.2021.07.011"},{"key":"2023092204350854000_2023.09.18.558302v1.44","doi-asserted-by":"publisher","DOI":"10.1016\/j.chom.2017.01.005"},{"key":"2023092204350854000_2023.09.18.558302v1.45","doi-asserted-by":"publisher","DOI":"10.1128\/JB.185.9.2731-2738.2003"},{"key":"2023092204350854000_2023.09.18.558302v1.46","doi-asserted-by":"publisher","DOI":"10.1111\/febs.12234"},{"key":"2023092204350854000_2023.09.18.558302v1.47","doi-asserted-by":"publisher","DOI":"10.1385\/1-59259-890-0:571"},{"key":"2023092204350854000_2023.09.18.558302v1.48","doi-asserted-by":"publisher","DOI":"10.1007\/s00792-011-0402-3"},{"key":"2023092204350854000_2023.09.18.558302v1.49","doi-asserted-by":"publisher","DOI":"10.1074\/jbc.M706996200"},{"key":"2023092204350854000_2023.09.18.558302v1.50","doi-asserted-by":"crossref","unstructured":"He, D. et al. Structural characterization of encapsulated ferritin provides insight into iron storage in bacterial nanocompartments. Elife 5(2016).","DOI":"10.7554\/eLife.18972"},{"key":"2023092204350854000_2023.09.18.558302v1.51","doi-asserted-by":"crossref","unstructured":"Suhanovsky, M.M. & Teschke, C.M . Nature\u2019s favorite building block: Deciphering folding and capsid assembly of proteins with the HK97-fold. Virology 479\u2013480, 487-97 (2015).","DOI":"10.1016\/j.virol.2015.02.055"},{"key":"2023092204350854000_2023.09.18.558302v1.52","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/gky309"},{"key":"2023092204350854000_2023.09.18.558302v1.53","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/gks363"},{"key":"2023092204350854000_2023.09.18.558302v1.54","doi-asserted-by":"crossref","first-page":"1388","DOI":"10.1021\/acs.biomac.2c01424","article-title":"Exploring the Extreme Acid Tolerance of a Dynamic Protein Nanocage","volume":"24","year":"2023","journal-title":"Biomacromolecules"},{"key":"2023092204350854000_2023.09.18.558302v1.55","doi-asserted-by":"publisher","DOI":"10.1126\/sciadv.abj4461"},{"key":"2023092204350854000_2023.09.18.558302v1.56","doi-asserted-by":"crossref","unstructured":"Kwon, S. , Andreas, M.P. & Giessen, T.W . Structure and heterogeneity of a highly cargo-loaded encapsulin shell. J Struct Biol 215, 108022 (2023).","DOI":"10.1016\/j.jsb.2023.108022"},{"key":"2023092204350854000_2023.09.18.558302v1.57","doi-asserted-by":"publisher","DOI":"10.1002\/pro.4582"},{"key":"2023092204350854000_2023.09.18.558302v1.58","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/gkw408"},{"key":"2023092204350854000_2023.09.18.558302v1.59","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/gkq399"},{"key":"2023092204350854000_2023.09.18.558302v1.60","doi-asserted-by":"publisher","DOI":"10.1128\/AEM.65.3.1029-1035.1999"},{"key":"2023092204350854000_2023.09.18.558302v1.61","doi-asserted-by":"publisher","DOI":"10.1007\/s00253-009-2173-7"},{"key":"2023092204350854000_2023.09.18.558302v1.62","doi-asserted-by":"publisher","DOI":"10.1038\/srep08245"},{"key":"2023092204350854000_2023.09.18.558302v1.63","doi-asserted-by":"publisher","DOI":"10.1007\/s00253-007-1261-9"},{"key":"2023092204350854000_2023.09.18.558302v1.64","doi-asserted-by":"publisher","DOI":"10.1007\/s00253-009-2369-x"},{"key":"2023092204350854000_2023.09.18.558302v1.65","doi-asserted-by":"crossref","first-page":"780","DOI":"10.1016\/j.chom.2020.09.005","article-title":"Epithelial-Derived Reactive Oxygen Species Enable AppBCX-Mediated Aerobic Respiration of Escherichia coli during Intestinal Inflammation","volume":"28","year":"2020","journal-title":"Cell Host Microbe"},{"key":"2023092204350854000_2023.09.18.558302v1.66","doi-asserted-by":"crossref","first-page":"789","DOI":"10.1016\/j.chom.2020.10.009","article-title":"Anaerobic Respiration of NOX1-Derived Hydrogen Peroxide Licenses Bacterial Growth at the Colonic Surface","volume":"28","year":"2020","journal-title":"Cell Host Microbe"},{"key":"2023092204350854000_2023.09.18.558302v1.67","doi-asserted-by":"publisher","DOI":"10.1038\/nmeth.4169"},{"key":"2023092204350854000_2023.09.18.558302v1.68","doi-asserted-by":"publisher","DOI":"10.1038\/nmeth.2019"},{"key":"2023092204350854000_2023.09.18.558302v1.69","doi-asserted-by":"publisher","DOI":"10.1107\/S0907444910007493"},{"key":"2023092204350854000_2023.09.18.558302v1.70","doi-asserted-by":"publisher","DOI":"10.1107\/S0907444909052925"},{"key":"2023092204350854000_2023.09.18.558302v1.71","doi-asserted-by":"publisher","DOI":"10.1002\/jcc.20084"},{"key":"2023092204350854000_2023.09.18.558302v1.72","doi-asserted-by":"publisher","DOI":"10.1002\/pro.3235"},{"key":"2023092204350854000_2023.09.18.558302v1.73","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/gkz303"},{"key":"2023092204350854000_2023.09.18.558302v1.74","doi-asserted-by":"crossref","first-page":"eabl7346","DOI":"10.1126\/sciadv.abl7346","article-title":"Pore structure controls stability and molecular flux in engineered protein cages","volume":"8","year":"2022","journal-title":"Sci Adv"},{"key":"2023092204350854000_2023.09.18.558302v1.75","doi-asserted-by":"publisher","DOI":"10.1038\/msb.2011.75"},{"key":"2023092204350854000_2023.09.18.558302v1.76","doi-asserted-by":"publisher","DOI":"10.1016\/j.jsb.2005.03.010"},{"key":"2023092204350854000_2023.09.18.558302v1.77","doi-asserted-by":"publisher","DOI":"10.1002\/PRO.3943"},{"key":"2023092204350854000_2023.09.18.558302v1.78","doi-asserted-by":"publisher","DOI":"10.1107\/s2059798319011471"},{"key":"2023092204350854000_2023.09.18.558302v1.79","doi-asserted-by":"crossref","first-page":"205","DOI":"10.1038\/s41592-022-01685-y","article-title":"AlphaFill: enriching AlphaFold models with ligands and cofactors","volume":"20","year":"2023","journal-title":"Nat Methods"}],"container-title":[],"original-title":[],"link":[{"URL":"https:\/\/syndication.highwire.org\/content\/doi\/10.1101\/2023.09.18.558302","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,1,15]],"date-time":"2026-01-15T05:15:06Z","timestamp":1768454106000},"score":1,"resource":{"primary":{"URL":"http:\/\/biorxiv.org\/lookup\/doi\/10.1101\/2023.09.18.558302"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2023,9,18]]},"references-count":79,"URL":"https:\/\/doi.org\/10.1101\/2023.09.18.558302","relation":{},"subject":[],"published":{"date-parts":[[2023,9,18]]},"subtype":"preprint"}}