{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,8,7]],"date-time":"2026-08-07T15:50:12Z","timestamp":1786117812971,"version":"build-2736575974"},"reference-count":46,"publisher":"American Society for Microbiology","issue":"8","license":[{"start":{"date-parts":[[2026,8,4]],"date-time":"2026-08-04T00:00:00Z","timestamp":1785801600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"},{"start":{"date-parts":[[2026,8,4]],"date-time":"2026-08-04T00:00:00Z","timestamp":1785801600000},"content-version":"tdm","delay-in-days":0,"URL":"https:\/\/journals.asm.org\/non-commercial-tdm-license"}],"funder":[{"DOI":"10.13039\/100009139","name":"Deutsches Zentrum f\u00fcr Infektionsforschung","doi-asserted-by":"publisher","award":["390838134"],"award-info":[{"award-number":["390838134"]}],"id":[{"id":"10.13039\/100009139","id-type":"DOI","asserted-by":"publisher"}]},{"name":"Medizinische Fakult\u00e4t, Eberhard Karls Universit\u00e4t T\u00fcbingen","award":["E.05.00849"],"award-info":[{"award-number":["E.05.00849"]}]}],"content-domain":{"domain":["journals.asm.org"],"crossmark-restriction":true},"short-container-title":["Microbiol Spectr"],"published-print":{"date-parts":[[2026,8,4]]},"abstract":"<jats:title>ABSTRACT<\/jats:title>\n                  <jats:sec>\n                    <jats:title\/>\n                    <jats:p>\n                      We previously described that a non-toxic ceftazidime-hexa-arginine conjugate (CTZ-R6) surpassed the antibacterial potency of ceftazidime (CTZ) against multi-resistant bacteria by up to 1,000-fold and effectively reduced the bacterial load in murine bacteremia. However, the mechanistic understanding of its resistance-overcoming activity remained elusive. Here, we report on the mode of action of CTZ-R6 in gram-positive and gram-negative bacteria. Against a panel of multi-resistant Enterobacteriaceae and non-fermenters, CTZ-R6 outperformed ceftazidime. The conjugate synergized with avibactam in the same manner as CTZ, but its MICs were much lower than those of CTZ. CTZ-R6 crossed the outer membrane of\n                      <jats:italic toggle=\"yes\">Escherichia coli<\/jats:italic>\n                      by self-promoted uptake and permeabilized this barrier to a fluorescent dye and otherwise ineffective antibiotics. Investigations in\n                      <jats:italic toggle=\"yes\">Bacillus subtilis<\/jats:italic>\n                      demonstrated that the electrochemical gradient at the cytoplasmic membrane remained high, but membrane topology was perturbed, and the peptidoglycan sacculus was damaged. Lysis kinetics were \u03b2-lactam-like. CTZ-R6 showed higher affinity than CTZ for penicillin-binding protein 2a (PBP2a) of\n                      <jats:italic toggle=\"yes\">B. subtilis<\/jats:italic>\n                      , lower affinity for PBP1a\/b, and an overall more balanced PBP affinity than CTZ. The covalent linkage between CTZ and the hexa-arginine moiety was essential for potency. In dissecting the contributions of the individual moieties, we found that R6C destabilizes the outer membrane, enabling porin-independent uptake. The peptide also interacts with the cytoplasmic membrane, facilitating CTZ access to the PBPs in\n                      <jats:italic toggle=\"yes\">B. subtilis<\/jats:italic>\n                      . More balanced targeting of PBPs and porin-independent entry is beneficial for overcoming established resistances. In conclusion, both components of the conjugate contribute to the increased antibacterial potency of CTZ-R6 in a balanced and synergistic manner.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>IMPORTANCE<\/jats:title>\n                    <jats:p>Given the global scale of antibacterial resistance, it becomes clear that numerous infections may eventually become untreatable as former, well-established treatment regimens fail. To prevent the emergence of a post-antibiotic era, strategies are being explored to extend the usefulness of clinically important antibiotics, including the synthesis of conjugates that improve the potency of established drugs. We recently demonstrated that attaching polycationic peptides to all four \u03b2-lactam subclasses improved MICs against multi-resistant gram-positive and gram-negative pathogens. The ceftazidime-hexa-arginine conjugate showed the best overall profile and improved the potency of ceftazidime against multi-resistant bacteria by up to 1,000-fold. However, the molecular mechanism of its resistance-overcoming activity remained elusive. Here, we describe which characteristics and capabilities of the peptide and the \u03b2-lactam allow them to synergize for increased potency. Mechanistic understanding is essential for further development of such conjugation approaches.<\/jats:p>\n                  <\/jats:sec>","DOI":"10.1128\/spectrum.00693-26","type":"journal-article","created":{"date-parts":[[2026,7,13]],"date-time":"2026-07-13T13:00:20Z","timestamp":1783947620000},"update-policy":"https:\/\/doi.org\/10.1128\/asmj-crossmark-policy-page","source":"Crossref","is-referenced-by-count":0,"title":["Synergistic mode of action of a ceftazidime-hexa-arginine conjugate with resistance-overcoming properties"],"prefix":"10.1128","volume":"14","author":[{"ORCID":"https:\/\/orcid.org\/0009-0004-1306-6620","authenticated-orcid":false,"given":"Hannah Isabell","family":"M\u00fcller","sequence":"first","affiliation":[{"id":[{"id":"https:\/\/ror.org\/03a1kwz48","id-type":"ROR","asserted-by":"publisher"}],"name":"Microbial Bioactive Compounds, Interfaculty Institute of Microbiology and Infection Medicine, University of T\u00fcbingen","place":["T\u00fcbingen, Germany"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Julia","family":"Werner","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/013czdx64","id-type":"ROR","asserted-by":"publisher"}],"name":"Department of Nuclear Medicine, University Hospital Heidelberg","place":["Heidelberg, Germany"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Julia","family":"Guther","sequence":"additional","affiliation":[{"name":"Institute of Medical Microbiology and Hygiene, University Hospital T\u00fcbingen","place":["T\u00fcbingen, Germany"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jan","family":"Straetener","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/03a1kwz48","id-type":"ROR","asserted-by":"publisher"}],"name":"Microbial Bioactive Compounds, Interfaculty Institute of Microbiology and Infection Medicine, University of T\u00fcbingen","place":["T\u00fcbingen, Germany"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Manuel Ben","family":"B\u00f6hmann","sequence":"additional","affiliation":[{"name":"Department of Pharmaceutical Technology, Institute of Pharmacy and Molecular Biotechnology, University of Heidelberg","place":["Heidelberg, Germany"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8616-5599","authenticated-orcid":true,"given":"Philipp","family":"Uhl","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/013czdx64","id-type":"ROR","asserted-by":"publisher"}],"name":"Department of Nuclear Medicine, University Hospital Heidelberg","place":["Heidelberg, Germany"]},{"name":"Department of Pharmaceutical Technology, Institute of Pharmacy and Molecular Biotechnology, University of Heidelberg","place":["Heidelberg, Germany"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9364-1832","authenticated-orcid":true,"given":"Heike","family":"Br\u00f6tz-Oesterhelt","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/03a1kwz48","id-type":"ROR","asserted-by":"publisher"}],"name":"Microbial Bioactive Compounds, Interfaculty Institute of Microbiology and Infection Medicine, University of T\u00fcbingen","place":["T\u00fcbingen, Germany"]},{"name":"Cluster of Excellence Controlling Microbes to Fight Infection","place":["T\u00fcbingen, Germany"]},{"id":[{"id":"https:\/\/ror.org\/028s4q594","id-type":"ROR","asserted-by":"publisher"}],"name":"German Center for Infection Research, Partner Site T\u00fcbingen","place":["T\u00fcbingen, Germany"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"235","reference":[{"key":"e_1_3_4_2_2","doi-asserted-by":"publisher","DOI":"10.1016\/S0140-6736(24)01867-1"},{"key":"e_1_3_4_3_2","doi-asserted-by":"publisher","DOI":"10.1179\/2047773215Y.0000000030"},{"key":"e_1_3_4_4_2","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pmed.1004264"},{"key":"e_1_3_4_5_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.drup.2023.101012"},{"key":"e_1_3_4_6_2","doi-asserted-by":"publisher","DOI":"10.1101\/cshperspect.a025247"},{"key":"e_1_3_4_7_2","doi-asserted-by":"publisher","DOI":"10.3390\/pathogens14040307"},{"key":"e_1_3_4_8_2","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.54.4.1133"},{"key":"e_1_3_4_9_2","doi-asserted-by":"publisher","DOI":"10.1101\/cshperspect.a025239"},{"key":"e_1_3_4_10_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.jmb.2019.04.002"},{"key":"e_1_3_4_11_2","doi-asserted-by":"publisher","DOI":"10.2165\/00003495-198530050-00002"},{"key":"e_1_3_4_12_2","doi-asserted-by":"publisher","DOI":"10.1002\/phar.1622"},{"key":"e_1_3_4_13_2","doi-asserted-by":"publisher","DOI":"10.1186\/s13756-025-01518-5"},{"key":"e_1_3_4_14_2","doi-asserted-by":"publisher","DOI":"10.2165\/11317030-000000000-00000"},{"key":"e_1_3_4_15_2","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pone.0251594"},{"key":"e_1_3_4_16_2","doi-asserted-by":"publisher","DOI":"10.1128\/AAC.30.1.1"},{"key":"e_1_3_4_17_2","doi-asserted-by":"publisher","DOI":"10.3390\/ph13060110"},{"key":"e_1_3_4_18_2","doi-asserted-by":"publisher","DOI":"10.1002\/anie.202002727"},{"key":"e_1_3_4_19_2","doi-asserted-by":"publisher","DOI":"10.1002\/advs.202411406"},{"key":"e_1_3_4_20_2","doi-asserted-by":"publisher","DOI":"10.4161\/viru.21282"},{"key":"e_1_3_4_21_2","doi-asserted-by":"publisher","DOI":"10.25646\/5916"},{"key":"e_1_3_4_22_2","unstructured":"The European Committee on Antimicrobial Susceptibility Testing. 2026. Breakpoint tables for interpretation of MICs and zone diameters. Available from: http:\/\/www.eucast.org"},{"key":"e_1_3_4_23_2","doi-asserted-by":"publisher","DOI":"10.1021\/acs.biomac.4c01706"},{"key":"e_1_3_4_24_2","doi-asserted-by":"publisher","DOI":"10.1074\/jbc.274.13.8405"},{"key":"e_1_3_4_25_2","doi-asserted-by":"publisher","DOI":"10.1128\/mBio.01659-18"},{"key":"e_1_3_4_26_2","doi-asserted-by":"publisher","DOI":"10.1128\/AEM.00586-07"},{"key":"e_1_3_4_27_2","doi-asserted-by":"publisher","DOI":"10.1038\/s42003-021-02137-7"},{"key":"e_1_3_4_28_2","doi-asserted-by":"publisher","DOI":"10.1177\/1087057107308161"},{"key":"e_1_3_4_29_2","doi-asserted-by":"publisher","DOI":"10.1021\/acsbiomedchemau.2c00007"},{"key":"e_1_3_4_30_2","doi-asserted-by":"publisher","DOI":"10.3390\/membranes10080181"},{"key":"e_1_3_4_31_2","doi-asserted-by":"publisher","DOI":"10.1038\/s42003-022-04035-y"},{"key":"e_1_3_4_32_2","doi-asserted-by":"publisher","DOI":"10.3390\/molecules24020249"},{"key":"e_1_3_4_33_2","doi-asserted-by":"publisher","DOI":"10.1155\/2015\/679109"},{"key":"e_1_3_4_34_2","doi-asserted-by":"publisher","DOI":"10.3390\/antibiotics14100958"},{"key":"e_1_3_4_35_2","doi-asserted-by":"publisher","DOI":"10.1002\/adtp.202400473"},{"key":"e_1_3_4_36_2","doi-asserted-by":"publisher","DOI":"10.1146\/annurev.mi.38.100184.001321"},{"key":"e_1_3_4_37_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.bmc.2016.06.044"},{"key":"e_1_3_4_38_2","doi-asserted-by":"publisher","DOI":"10.3389\/fmicb.2020.593215"},{"key":"e_1_3_4_39_2","doi-asserted-by":"publisher","DOI":"10.1007\/978-3-030-16373-0_5"},{"key":"e_1_3_4_40_2","doi-asserted-by":"publisher","DOI":"10.3389\/fmed.2021.677720"},{"key":"e_1_3_4_41_2","unstructured":"CLSI. 2022. Performance standards for antimicrobial susceptibility testing. 32nd ed. Clinical and Laboratory Standards Institute"},{"key":"e_1_3_4_42_2","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.1319900111"},{"key":"e_1_3_4_43_2","doi-asserted-by":"publisher","DOI":"10.1099\/00221287-139-9-2041"},{"key":"e_1_3_4_44_2","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.1110385108"},{"key":"e_1_3_4_45_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.bbamem.2018.01.011"},{"key":"e_1_3_4_46_2","doi-asserted-by":"publisher","DOI":"10.3389\/fmicb.2018.00019"},{"key":"e_1_3_4_47_2","doi-asserted-by":"publisher","DOI":"10.1093\/jac\/29.1.27"}],"container-title":["Microbiology Spectrum"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/journals.asm.org\/doi\/pdf\/10.1128\/spectrum.00693-26","content-type":"application\/pdf","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/journals.asm.org\/doi\/pdf\/10.1128\/spectrum.00693-26","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,8,6]],"date-time":"2026-08-06T14:41:56Z","timestamp":1786027316000},"score":1,"resource":{"primary":{"URL":"https:\/\/journals.asm.org\/doi\/10.1128\/spectrum.00693-26"}},"subtitle":[],"editor":[{"given":"Christine","family":"Josenhans","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"editor"}]}],"short-title":[],"issued":{"date-parts":[[2026,8,4]]},"references-count":46,"journal-issue":{"issue":"8","published-print":{"date-parts":[[2026,8,4]]}},"alternative-id":["10.1128\/spectrum.00693-26"],"URL":"https:\/\/doi.org\/10.1128\/spectrum.00693-26","relation":{},"ISSN":["2165-0497"],"issn-type":[{"value":"2165-0497","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,8,4]]},"assertion":[{"value":"2026-02-27","order":0,"name":"received","label":"Received","group":{"name":"publication_history","label":"Publication History"}},{"value":"2026-05-29","order":2,"name":"accepted","label":"Accepted","group":{"name":"publication_history","label":"Publication History"}},{"value":"2026-07-13","order":3,"name":"published","label":"Published","group":{"name":"publication_history","label":"Publication History"}}],"article-number":"e00693-26"}}