title: Acyl carrier protein is a bacterial cytoplasmic target of cationic antimicrobial peptide LL-37.
abstract: 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.
authors.affiliation: School of Systems Biology and the National Center for Biodefense and Infectious Diseases, George Mason University, Manassas, VA 20110, U.S.A.
authors.forename: Myung-Chul
authors.initials: MC
authors.lastname: Chung
authors.affiliation: School of Systems Biology and the National Center for Biodefense and Infectious Diseases, George Mason University, Manassas, VA 20110, U.S.A.
authors.forename: Scott N
authors.initials: SN
authors.lastname: Dean
authors.affiliation: School of Systems Biology and the National Center for Biodefense and Infectious Diseases, George Mason University, Manassas, VA 20110, U.S.A. mvanhoek@gmu.edu.
authors.forename: Monique L
authors.initials: ML
authors.lastname: van Hoek
mesh_terms: D000213:Acyl Carrier Protein / Q000378:metabolism*; D000595:Amino Acid Sequence; D000900:Anti-Bacterial Agents / Q000378:metabolism* / Q000494:pharmacology; D023181:Antimicrobial Cationic Peptides / Q000378:metabolism* / Q000494:pharmacology; D001426:Bacterial Proteins / Q000378:metabolism*; D003593:Cytoplasm / Q000378:metabolism; D005227:Fatty Acids / Q000378:metabolism; D005603:Francisella / Q000187:drug effects / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011485:Protein Binding; D054804:Cathelicidins
chemical_list: D000213:Acyl Carrier Protein; D000900:Anti-Bacterial Agents; D023181:Antimicrobial Cationic Peptides; D001426:Bacterial Proteins; D005227:Fatty Acids; D054804:Cathelicidins
keywords: Francisella; acyl carrier protein; antimicrobial peptide; cathelicidin; drug target