biostudies-literatureBello-Madruga Ra node of the Spanish Large-Scale National FacilityMinisterio de Ciencia e Innovación (MCIN)NMR experiments were performed in the “Manuel Rico” NMR laboratory (LMR) of the Spanish National Research CouncilMinisterio de Ciencia e Innovaciónthe Departament de Recerca i Universitats de la Generalitat de CatalunyaEuropean Society of Clinical Microbiology and Infectious Diseases'la Caixa' Foundation ('la Caixa')European Society of Clinical Microbiology and Infectious Diseases (ESCMID)'la Caixa' Foundation889-910https://www.ebi.ac.uk/biostudies/studies/S-EPMC12223310biostudies-literatureUnknown21(7)Glycosaminoglycan (GAG)-binding proteins regulating essential processes such as cell growth and migration are essential for cell homeostasis. As both GAGs and the lipid A disaccharide core of Gram-negative bacteria contain negatively charged disaccharide units, we hypothesized that GAG-binding proteins could also recognize LPS and enclose cryptic antibiotic motifs. Here, we report novel antimicrobial peptides (AMPs) derived from heparin-binding proteins (HBPs), with specific activity against Gram-negative bacteria and high LPS binding. We used computational tools to locate antimicrobial regions in 82% of HBPs, most of those colocalizing with putative heparin-binding sites. To validate these results, we synthesized five candidates [HBP-1-5] that showed remarkable activity against Gram-negatMolecular systems biologyMining the heparinome for cryptic antimicrobial peptides that selectively kill Gram-negative bacteria.PMC12223310ESCMID2022PID2020-113184RB-C22PRTR-C17.I1PDC2021-121544-I00 funded by MCIN/AEI /10.13039/501100011033 and European Union Next GenerationEU/ PHR17-00409PRE2021-097678María de Maeztu network of Units of ExcellencePRE2018-083243PID2020-112821GB-I00 funded by MCIN/AEI /10.13039/501100011033PID2020-114627RB-I00 funded by MCIN/AEI /10.13039/5011000110332023PROD00021Gomez JBello-Madruga RTorrent MValle JComas LGonzalez-Lopez JJJimenez MASandin DLarrosa MNAndreu DfalseMining the heparinome for cryptic antimicrobial peptides that selectively kill Gram-negative bacteria.Glycosaminoglycan (GAG)-binding proteins regulating essential processes such as cell growth and migration are essential for cell homeostasis. As both GAGs and the lipid A disaccharide core of Gram-negative bacteria contain negatively charged disaccharide units, we hypothesized that GAG-binding proteins could also recognize LPS and enclose cryptic antibiotic motifs. Here, we report novel antimicrobial peptides (AMPs) derived from heparin-binding proteins (HBPs), with specific activity against Gram-negative bacteria and high LPS binding. We used computational tools to locate antimicrobial regions in 82% of HBPs, most of those colocalizing with putative heparin-binding sites. To validate these results, we synthesized five candidates [HBP-1-5] that showed remarkable activity against Gram-negat2025-01-01T00:00:00Z2025 Jul2026-06-03T01:43:23.393Z2026-05-28T03:05:21.801ZS-EPMC122233104041038210.1038/s44320-025-00120-6