Scientists discovered that the human antimicrobial peptide LL‑37 can slip inside bacteria and latch onto a protein called AcpP, which is involved in making fatty acids. This binding changes the bacteria's fat makeup and makes them more vulnerable to LL‑37, especially when another peptide, SMAP‑29, is present. The finding shows LL‑37 works not just by punching holes in bacterial membranes but also by targeting internal machinery.

In addition to membrane disruption, the cathelicidin antimicrobial peptide (AMP) LL-37 translocates through the bacterial inner membrane to target intracellular molecules. The present study aims to identify an alternate mechanism and a cytoplasmic target of LL-37 in Francisella. LL-37 binding proteins from Francisella novicida U112 bacterial lysates were precipitated by using biotinylated LL-37 (B-LL-37) and NeutrAvidin-agarose beads. Bound proteins were identified by LC-MS/MS, validated and characterized by bead pull-down assays and differential scanning fluorimetry (DSF). The cationic AMP (CAMP) LL-37 was able to interact with Francisella cytoplasmic acyl carrier protein (AcpP; FTN1340/FTT1376). Further study confirmed that LL-37 peptide could bind to AcpP and that the sheep cathelicidin SMAP-29 (Sheep Myeloid Antimicrobial Peptide 29) further increased LL-37 binding to AcpP, suggesting a synergistic effect of SMAP-29 on the binding. LL-37 could also bind to both AcpP of Escherichia coli and Bacillus anthracis, implying a mechanism of broad action of LL-37-AcpP binding. Overexpression of the acpP gene in F. novicida led to an increase in LL-37 susceptibility. LL-37 binding to AcpP changed the fatty acid composition profiles. Taken together, we identified a novel cytoplasmic target of LL-37 in Francisella, suggesting a mechanism of action of this peptide beyond membrane permeabilization. Our findings highlight a novel mechanism of antimicrobial activity of this peptide and document a previously unexplored target of α-helical CAMPs.