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Novel treatments are needed, but are difficult to develop for Gram-negative species due to the need to traverse the outer membrane to reach targets beneath. A promising solution is found in natural antibiotics which bind outer membrane receptors and co-opt them for import. Exploring this mechanism may open avenues for antibiotic development. An underappreciated class of natural antibiotics are microcins - small antimicrobial proteins secreted by certain bacteria during inter-species competition. Microcins bind outer-membrane receptors of prey species for passage into the periplasm. They have potent activity, bind specific targets, and can control pathobiont expansion and colonization. One microcin, MccV, utilizes the <jats:italic>E. coli<\/jats:italic> colicin Ia receptor, Cir, for import. Here, we report the first high-resolution structure of the Cir\/MccV complex by Cryo-EM, revealing an interaction centered on an electropositive cavity within the Cir extracellular loops. We also report the affinity of MccV for Cir. Lastly, we mutagenized interacting residues and identified key contacts critical to MccV binding, import, and bacteriolysis. Future efforts may help disentangle the mechanisms of microcin killing and will assess relationships between other microcins and their targets to better understand the potential for microcins to be used as antibacterial drugs.<\/jats:p>","DOI":"10.1038\/s42003-025-08846-7","type":"journal-article","created":{"date-parts":[[2025,10,9]],"date-time":"2025-10-09T11:53:10Z","timestamp":1760010790000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":4,"title":["Structural insights into Cir-mediated killing by the antimicrobial protein Microcin V"],"prefix":"10.1038","volume":"8","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-8750-6279","authenticated-orcid":false,"given":"Stavros A.","family":"Maurakis","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Angela C.","family":"O\u2019Donnell","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Istvan","family":"Botos","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4880-4959","authenticated-orcid":false,"given":"Rodolfo","family":"Ghirlando","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2046-0341","authenticated-orcid":false,"given":"Bryan W.","family":"Davies","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9657-7119","authenticated-orcid":false,"given":"Susan K.","family":"Buchanan","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"297","published-online":{"date-parts":[[2025,10,9]]},"reference":[{"key":"8846_CR1","unstructured":"Global burden of bacterial antimicrobial resistance in 2019: a systematic analysis. 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