{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,28]],"date-time":"2026-10-28T05:36:52Z","timestamp":1793165812879,"version":"4.5.0"},"reference-count":43,"publisher":"Wiley","issue":"10","license":[{"start":{"date-parts":[[2026,10,16]],"date-time":"2026-10-16T00:00:00Z","timestamp":1792108800000},"content-version":"vor","delay-in-days":15,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"},{"start":{"date-parts":[[2026,10,1]],"date-time":"2026-10-01T00:00:00Z","timestamp":1790812800000},"content-version":"tdm","delay-in-days":0,"URL":"http:\/\/doi.wiley.com\/10.1002\/tdm_license_1.1"}],"funder":[{"DOI":"10.13039\/501100024250","name":"Israel Innovation Authority","doi-asserted-by":"publisher","award":["81260"],"award-info":[{"award-number":["81260"]}],"id":[{"id":"10.13039\/501100024250","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100024250","name":"Israel Innovation Authority","doi-asserted-by":"publisher","award":["81259"],"award-info":[{"award-number":["81259"]}],"id":[{"id":"10.13039\/501100024250","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["enviromicro-journals.onlinelibrary.wiley.com"],"crossmark-restriction":true},"short-container-title":["Microbial Biotechnology"],"published-print":{"date-parts":[[2026,10]]},"abstract":"<jats:title>ABSTRACT<\/jats:title>\n                  <jats:p>Bacterial protein toxins constitute a vast, largely untapped reservoir of potent bioactive molecules for antimicrobial discovery and biotechnology. However, their intrinsic toxicity often prevents recombinant expression in microbial hosts, limiting biochemical and functional characterization. Here, we present a modular platform for the recombinant production of highly toxic bacterial proteins based on covalent fusion of polymorphic toxins (PTs) to their cognate immunity proteins (PIMs), enabling transient toxin neutralization during expression. This strategy enabled recombinant production and purification of three previously refractory toxin families, PT7, PT8 and PT9. We further streamlined the workflow by introducing intracellular protease\u2010mediated cleavage of the toxin\u2010immunity fusion, reducing purification from four steps to two and increasing the recovered yield of PT9 from approximately 1\u20132\u2009mg\/L to ~4\u2009mg\/L. Extending the approach to PT37 further showed that computational redesign of the immunity protein and fusion architecture can enable production when the native fusion is insufficient. Purified PT7 and PT8 retained nuclease activity following denaturation and refolding, while refolded PT9 was structurally validated by X\u2010ray crystallography, demonstrating recovery of functionally and structurally competent proteins. Together, these findings establish a modular strategy for recombinant production of highly toxic bacterial proteins and facilitate their biochemical and functional characterization.<\/jats:p>","DOI":"10.1111\/1751-7915.70453","type":"journal-article","created":{"date-parts":[[2026,10,17]],"date-time":"2026-10-17T06:10:22Z","timestamp":1792217422000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Modular Platform for Recombinant Production of Highly Toxic Bacterial Proteins"],"prefix":"10.1111","volume":"19","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-5283-9532","authenticated-orcid":false,"given":"Rina","family":"Fraenkel","sequence":"first","affiliation":[{"name":"Department of Biological Chemistry, Alexander Silberman Institute of Life Sciences, Faculty of Mathematics &amp; Science The Hebrew University of Jerusalem  Jerusalem Israel"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3030-7666","authenticated-orcid":false,"given":"Inbar","family":"Cahana","sequence":"additional","affiliation":[{"name":"Department of Biological Chemistry, Alexander Silberman Institute of Life Sciences, Faculty of Mathematics &amp; Science The Hebrew University of Jerusalem  Jerusalem Israel"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0004-6565-2701","authenticated-orcid":false,"given":"Tomer","family":"Sivan","sequence":"additional","affiliation":[{"name":"Department of Biological Chemistry, Alexander Silberman Institute of Life Sciences, Faculty of Mathematics &amp; Science The Hebrew University of Jerusalem  Jerusalem Israel"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0001-8624-4000","authenticated-orcid":false,"given":"Tal","family":"Fisher","sequence":"additional","affiliation":[{"name":"Department of Biological Chemistry, Alexander Silberman Institute of Life Sciences, Faculty of Mathematics &amp; Science The Hebrew University of Jerusalem  Jerusalem Israel"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0008-3839-3782","authenticated-orcid":false,"given":"Hen","family":"Nadav","sequence":"additional","affiliation":[{"name":"Department of Biological Chemistry, Alexander Silberman Institute of Life Sciences, Faculty of Mathematics &amp; Science The Hebrew University of Jerusalem  Jerusalem Israel"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0001-0161-3074","authenticated-orcid":false,"given":"Shani","family":"Bruchim","sequence":"additional","affiliation":[{"name":"Department of Biological Chemistry, Alexander Silberman Institute of Life Sciences, Faculty of Mathematics &amp; Science The Hebrew University of Jerusalem  Jerusalem Israel"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0003-3895-3990","authenticated-orcid":false,"given":"Jenna","family":"Weisz","sequence":"additional","affiliation":[{"name":"Department of Biological Chemistry, Alexander Silberman Institute of Life Sciences, Faculty of Mathematics &amp; Science The Hebrew University of Jerusalem  Jerusalem Israel"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-7885-4722","authenticated-orcid":false,"given":"Noam","family":"Deouell","sequence":"additional","affiliation":[{"name":"Department of Biological Chemistry, Alexander Silberman Institute of Life Sciences, Faculty of Mathematics &amp; Science The Hebrew University of Jerusalem  Jerusalem Israel"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0000-0255-2616","authenticated-orcid":false,"given":"Shani","family":"Cheskis","sequence":"additional","affiliation":[{"name":"The Department of Plant Pathology and Microbiology, Institute of Environmental Sciences, Robert H. 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