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We have developed novel AMP candidate (peptide 3.1) based on indolicidin, among the shortest naturally occurring AMP. The antimicrobial activity of this peptide is demonstrated by the minimum inhibitory concentration, while the hemolysis tests and MTT assay indicate its low cytotoxicity. In optical diffraction tomography, red blood cells treated with peptide 3.1 showed no discernible effects, in contrast to indolicidin. However, peptide 3.1 did induce cell lysis in E. coli, leading to a reduced potential for the development of antibiotic resistance. To investigate the mechanism underlying membrane selectivity, the structure of peptide 3.1 was analyzed using nuclear magnetic resonance spectroscopy and molecular dynamics simulations. Peptide 3.1 is structured with an increased distinction between hydrophobic and charged residues and remained in close proximity to the eukaryotic membrane. On the other hand, peptide 3.1 exhibited a disordered conformation when approaching the prokaryotic membrane, similar to indolicidin, leading to its penetration into the membrane. Consequently, it appears that the amphipathicity and structural rigidity of peptide 3.1 contribute to its membrane selectivity. In conclusion, this study may lead to the development of Peptide 3.1, a promising commercial candidate based on its low cost to produce and low cytotoxicity. We have also shed light on the mechanism of action of AMP, which exhibits selective toxicity to bacteria while not damaging eukaryotic cells.<\/jats:p>","DOI":"10.1002\/biof.2145","type":"journal-article","created":{"date-parts":[[2024,11,21]],"date-time":"2024-11-21T07:29:26Z","timestamp":1732174166000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":3,"title":["Insights into an indolicidin\u2010derived low\u2010toxic anti\u2010microbial peptide's efficacy against bacterial cells while preserving eukaryotic cell viability"],"prefix":"10.1002","volume":"51","author":[{"given":"Jihyun","family":"Kim","sequence":"first","affiliation":[{"name":"Department of New Biology DGIST  Daegu Republic of Korea"},{"name":"Repill Inc.  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Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Young Ho","family":"Ko","sequence":"additional","affiliation":[{"name":"Center for Self\u2010Assembly and Complexity, Institute for Basic Science  Pohang Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-6776-2451","authenticated-orcid":false,"given":"Jin Hae","family":"Kim","sequence":"additional","affiliation":[{"name":"Department of New Biology DGIST  Daegu Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Su Il","family":"In","sequence":"additional","affiliation":[{"name":"Department of Energy Science and Engineering DGIST  Daegu Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3570-7802","authenticated-orcid":false,"given":"Chang Hoon","family":"Nam","sequence":"additional","affiliation":[{"name":"Department of New Biology DGIST  Daegu Republic of 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