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The bacterial killing mechanism of chemokines and the biochemical properties that define which members of the chemokine superfamily are antimicrobial remain poorly understood. We report that the antimicrobial activity of chemokines is defined by their ability to bind phosphatidylglycerol and cardiolipin, two anionic phospholipids commonly found in the bacterial plasma membrane. We show that only chemokines able to bind these two phospholipids kill bacteria and that they exert rapid bacteriostatic and bactericidal effects with a higher potency than the antimicrobial peptide \u03b2-defensin 3. Both biochemical and genetic interference with the chemokine-cardiolipin interaction impaired microbial growth arrest, bacterial killing, and membrane disruption by chemokines. Moreover, unlike conventional antibiotics,\n                    <jats:italic toggle=\"yes\">Escherichia coli<\/jats:italic>\n                    failed to develop resistance when placed under increasing antimicrobial chemokine pressure in vitro. Thus, we have identified cardiolipin and phosphatidylglycerol as binding partners for chemokines responsible for chemokine antimicrobial action.\n                  <\/jats:p>","DOI":"10.1126\/sciadv.ads2675","type":"journal-article","created":{"date-parts":[[2025,6,6]],"date-time":"2025-06-06T13:58:24Z","timestamp":1749218304000},"update-policy":"https:\/\/doi.org\/10.34133\/aaas_crossmark","source":"Crossref","is-referenced-by-count":8,"title":["Chemokines kill bacteria without triggering antimicrobial resistance by binding anionic phospholipids"],"prefix":"10.1126","volume":"11","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-7756-9831","authenticated-orcid":true,"given":"Sergio M.","family":"Pontejo","sequence":"first","affiliation":[{"name":"Laboratory of Molecular Immunology, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0008-3078-3487","authenticated-orcid":true,"given":"Sophia","family":"Martinez","sequence":"additional","affiliation":[{"name":"Laboratory of Molecular Immunology, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0009-0761-9205","authenticated-orcid":true,"given":"Allison","family":"Zhao","sequence":"additional","affiliation":[{"name":"Laboratory of Molecular Immunology, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0006-7888-7512","authenticated-orcid":true,"given":"Kevin","family":"Barnes","sequence":"additional","affiliation":[{"name":"Structural Biology Section, Research Technologies Branch, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2129-0775","authenticated-orcid":true,"given":"Jaime","family":"de Anda","sequence":"additional","affiliation":[{"name":"Department of Bioengineering, University of California, Los Angeles, Los Angeles, CA 90095, USA."},{"name":"Department of Chemistry and Biochemistry, University of California, Los Angeles, Los Angeles, CA 90095, USA."},{"name":"Department of Microbiology, Immunology &amp; Molecular Genetics, University of California Los Angeles, Los Angeles, CA 90095, USA."},{"name":"California NanoSystems Institute, University of California Los Angeles, Los Angeles, CA 90095, USA."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-6576-2426","authenticated-orcid":true,"given":"Haleh","family":"Alimohamadi","sequence":"additional","affiliation":[{"name":"Department of Bioengineering, University of California, Los Angeles, Los Angeles, CA 90095, USA."},{"name":"Department of Chemistry and Biochemistry, University of California, Los Angeles, Los Angeles, CA 90095, USA."},{"name":"Department of Microbiology, Immunology &amp; Molecular Genetics, University of California Los Angeles, Los Angeles, CA 90095, USA."},{"name":"California NanoSystems Institute, University of California Los Angeles, Los Angeles, CA 90095, USA."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5144-2552","authenticated-orcid":true,"given":"Ernest Y.","family":"Lee","sequence":"additional","affiliation":[{"name":"Department of Bioengineering, University of California, Los Angeles, Los Angeles, CA 90095, USA."},{"name":"Department of Chemistry and Biochemistry, University of California, Los Angeles, Los Angeles, CA 90095, USA."},{"name":"Department of Microbiology, Immunology &amp; Molecular Genetics, University of California Los Angeles, Los Angeles, CA 90095, USA."},{"name":"California NanoSystems Institute, University of California Los Angeles, Los Angeles, CA 90095, USA."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Acacia F.","family":"Dishman","sequence":"additional","affiliation":[{"name":"Department of Biochemistry, Medical College of Wisconsin, Milwaukee, WI 53226, USA."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-6681-5179","authenticated-orcid":true,"given":"Brian F.","family":"Volkman","sequence":"additional","affiliation":[{"name":"Department of Biochemistry, Medical College of Wisconsin, Milwaukee, WI 53226, USA."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0893-6383","authenticated-orcid":true,"given":"Gerard C. 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