{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,4]],"date-time":"2026-09-04T19:12:44Z","timestamp":1788549164218,"version":"build-2803163510"},"reference-count":66,"publisher":"Wiley","issue":"8","license":[{"start":{"date-parts":[[2026,8,14]],"date-time":"2026-08-14T00:00:00Z","timestamp":1786665600000},"content-version":"vor","delay-in-days":13,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"},{"start":{"date-parts":[[2026,8,1]],"date-time":"2026-08-01T00:00:00Z","timestamp":1785542400000},"content-version":"tdm","delay-in-days":0,"URL":"http:\/\/doi.wiley.com\/10.1002\/tdm_license_1.1"}],"funder":[{"DOI":"10.13039\/501100000268","name":"Biotechnology and Biological Sciences Research Council","doi-asserted-by":"publisher","award":["BB\/T008768\/1"],"award-info":[{"award-number":["BB\/T008768\/1"]}],"id":[{"id":"10.13039\/501100000268","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100000268","name":"Biotechnology and Biological Sciences Research Council","doi-asserted-by":"publisher","award":["BB\/X007448\/1"],"award-info":[{"award-number":["BB\/X007448\/1"]}],"id":[{"id":"10.13039\/501100000268","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["J of Extracellular Vesicle"],"published-print":{"date-parts":[[2026,8]]},"abstract":"<jats:title>ABSTRACT<\/jats:title>\n                  <jats:p>\n                    Bacterial outer membrane vesicles (OMVs) are nano\u2010sized, spherical structures released by Gram\u2010negative bacteria that play diverse roles in bacterial physiology, including communication, nutrient acquisition and host interactions. These vesicles bud from the bacterial outer membrane and contain lipopolysaccharides, periplasmic proteins, nucleotides and other biomolecules. The vesicle nucleating peptide (VNp) is a short peptide tag that, when fused to the amino terminus of a protein of interest, promotes the formation of bespoke recombinant extracellular vesicles (EVs) in\n                    <jats:italic>Escherichia coli<\/jats:italic>\n                    , enabling efficient production and simplified purification of recombinant proteins. Here, we characterise VNp\u2010induced extracellular vesicles (VNp\u2010EVs) and compare their composition and organisation with OMVs produced from\n                    <jats:italic>E. coli<\/jats:italic>\n                    expressing a periplasmic targeting fusion. While both vesicle types possess a single outer membrane\u2010derived lipid bilayer, recombinant protein is highly enriched within the VNp\u2010EVs compared to OMVs containing the periplasm targeting ssDsbA\u2010fusion protein. VNp\u2010fusions and the periplasm\u2010targeted recombinant protein localise to distinct vesicle populations, with VNp\u2010fusions showing markedly higher luminal concentrations and relative vesicular abundance, compared to the vesicles containing a periplasmic targeted fusion protein. OmpX co\u2010expression further enriched the VNp\u2010fusion content of vesicles, further enhancing yield. The VNp\u2010vesicle lumen is an oxidising environment, thus supports formation of inter\u2010 and intra\u2010molecular disulfide bonds within encapsulated proteins. Overall, VNp\u2010EVs represent a distinct class of recombinant EVs that offer a simple and efficient route for producing and purifying concentrated, correctly folded recombinant proteins, expanding the utility of bacterial vesicle systems for biotechnological applications.\n                  <\/jats:p>","DOI":"10.1002\/jev2.70354","type":"journal-article","created":{"date-parts":[[2026,8,14]],"date-time":"2026-08-14T17:46:27Z","timestamp":1786729587000},"source":"Crossref","is-referenced-by-count":0,"title":["Vesicle Nucleation Peptide Fusion Induced Extracellular Vesicles Are Distinct From\n                    <i>Escherichia coli<\/i>\n                    Outer Membrane Vesicles, and Provide an Enhanced Platform for Protein Production and Purification"],"prefix":"10.1002","volume":"15","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-0190-8558","authenticated-orcid":false,"given":"Bree R.","family":"Streather","sequence":"first","affiliation":[{"name":"School of Biosciences University of Kent  Canterbury Kent UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2587-0574","authenticated-orcid":false,"given":"Tara A.","family":"Eastwood","sequence":"additional","affiliation":[{"name":"School of Biosciences University of Kent  Canterbury Kent UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7628-1978","authenticated-orcid":false,"given":"Karen","family":"Baker","sequence":"additional","affiliation":[{"name":"School of Biosciences University of Kent  Canterbury Kent UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8925-6739","authenticated-orcid":false,"given":"Mingzhi","family":"Liang","sequence":"additional","affiliation":[{"name":"School of Biosciences University of Kent  Canterbury Kent UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5641-6003","authenticated-orcid":false,"given":"Alexandra E.","family":"Bailie","sequence":"additional","affiliation":[{"name":"Central Laser Facility, Research Complex at Harwell Science and Technology Facilities Council, Rutherford Appleton Laboratory  Didcot Oxford UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8388-2624","authenticated-orcid":false,"given":"Tijn T.","family":"van der Velden","sequence":"additional","affiliation":[{"name":"Leiden Institute of Chemistry University of Leiden  Leiden the Netherlands"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Lars J. C.","family":"Jeuken","sequence":"additional","affiliation":[{"name":"Leiden Institute of Chemistry University of Leiden  Leiden the Netherlands"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3268-9303","authenticated-orcid":false,"given":"Stanley W.","family":"Botchway","sequence":"additional","affiliation":[{"name":"Central Laser Facility, Research Complex at Harwell Science and Technology Facilities Council, Rutherford Appleton Laboratory  Didcot Oxford UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9141-5217","authenticated-orcid":false,"given":"Lin","family":"Wang","sequence":"additional","affiliation":[{"name":"Central Laser Facility, Research Complex at Harwell Science and Technology Facilities Council, Rutherford Appleton Laboratory  Didcot Oxford UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2502-5274","authenticated-orcid":false,"given":"Daniel P.","family":"Mulvihill","sequence":"additional","affiliation":[{"name":"School of Biosciences University of Kent  Canterbury Kent UK"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"311","published-online":{"date-parts":[[2026,8,14]]},"reference":[{"key":"e_1_2_9_2_1","doi-asserted-by":"publisher","DOI":"10.1016\/0022-1759(90)90029-U"},{"key":"e_1_2_9_3_1","doi-asserted-by":"publisher","DOI":"10.1098\/rsob.240229"},{"key":"e_1_2_9_4_1","doi-asserted-by":"publisher","DOI":"10.1242\/jcs.190124"},{"key":"e_1_2_9_5_1","doi-asserted-by":"publisher","DOI":"10.1002\/cpz1.70255"},{"key":"e_1_2_9_6_1","doi-asserted-by":"publisher","DOI":"10.1016\/0092-8674(91)90532-4"},{"key":"e_1_2_9_7_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.ejca.2011.11.025"},{"key":"e_1_2_9_8_1","doi-asserted-by":"publisher","DOI":"10.1042\/bj0960567"},{"key":"e_1_2_9_9_1","doi-asserted-by":"publisher","DOI":"10.1002\/jemt.20548"},{"key":"e_1_2_9_10_1","doi-asserted-by":"publisher","DOI":"10.1111\/jmi.12218"},{"key":"e_1_2_9_11_1","doi-asserted-by":"publisher","DOI":"10.1002\/cm.22026"},{"key":"e_1_2_9_12_1","doi-asserted-by":"publisher","DOI":"10.1038\/s41598-022-26880-x"},{"key":"e_1_2_9_13_1","doi-asserted-by":"publisher","DOI":"10.1111\/j.1574-6976.2012.00327.x"},{"key":"e_1_2_9_14_1","doi-asserted-by":"publisher","DOI":"10.1111\/j.1365-2958.2009.06731.x"},{"key":"e_1_2_9_15_1","doi-asserted-by":"publisher","DOI":"10.1128\/jb.178.3.768-773.1996"},{"key":"e_1_2_9_16_1","doi-asserted-by":"publisher","DOI":"10.1128\/JB.00315-11"},{"key":"e_1_2_9_17_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.crmeth.2023.100396"},{"key":"e_1_2_9_18_1","doi-asserted-by":"publisher","DOI":"10.1099\/mic.0.060343-0"},{"key":"e_1_2_9_19_1","doi-asserted-by":"publisher","DOI":"10.1002\/jev2.70150"},{"key":"e_1_2_9_20_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.vaccine.2025.126899"},{"key":"e_1_2_9_21_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.micpath.2022.105965"},{"key":"e_1_2_9_22_1","doi-asserted-by":"publisher","DOI":"10.1002\/elps.200390069"},{"key":"e_1_2_9_23_1","doi-asserted-by":"publisher","DOI":"10.1074\/jbc.M110.185744"},{"key":"e_1_2_9_24_1","doi-asserted-by":"publisher","DOI":"10.1016\/0014-5793(93)80807-7"},{"key":"e_1_2_9_25_1","doi-asserted-by":"publisher","DOI":"10.1080\/20013078.2019.1632099"},{"key":"e_1_2_9_26_1","doi-asserted-by":"publisher","DOI":"10.1074\/jbc.REV120.011473"},{"key":"e_1_2_9_27_1","doi-asserted-by":"publisher","DOI":"10.1074\/jbc.275.17.12489"},{"key":"e_1_2_9_28_1","doi-asserted-by":"publisher","DOI":"10.1002\/bies.201700003"},{"key":"e_1_2_9_29_1","doi-asserted-by":"publisher","DOI":"10.1128\/JB.177.14.3998-4008.1995"},{"key":"e_1_2_9_30_1","doi-asserted-by":"publisher","DOI":"10.1074\/jbc.M307628200"},{"key":"e_1_2_9_31_1","doi-asserted-by":"publisher","DOI":"10.1128\/AEM.65.5.1843-1848.1999"},{"key":"e_1_2_9_32_1","doi-asserted-by":"publisher","DOI":"10.1038\/nmeth.4344"},{"key":"e_1_2_9_33_1","doi-asserted-by":"publisher","DOI":"10.3389\/fmicb.2018.02268"},{"key":"e_1_2_9_34_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.ces.2024.120809"},{"key":"e_1_2_9_35_1","doi-asserted-by":"publisher","DOI":"10.1016\/S0009-9120(97)00049-0"},{"key":"e_1_2_9_36_1","doi-asserted-by":"publisher","DOI":"10.1128\/jb.137.2.1031-1034.1979"},{"issue":"3","key":"e_1_2_9_37_1","first-page":"388","article-title":"Methods of Endotoxin Removal From Biological Preparations: A Review","volume":"10","author":"Magalhaes P. 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