Structural remodeling and oligomerization of human cathelicidin on membranes suggest fibril-like structures as active species

Abstract

Antimicrobial peptides as part of the mammalian innate immune system target and remove major bacterial pathogens, often through irreversible damage of their cellular membranes. To explore the mechanism by which the important cathelicidin peptide LL-37 of the human innate immune system interacts with membranes, we performed biochemical, biophysical and structural studies. The crystal structure of LL-37 displays dimers of anti-parallel helices and the formation of amphipathic surfaces. Peptide-detergent interactions introduce remodeling of this structure after occupation of defined hydrophobic sites at the dimer interface. Furthermore, hydrophobic nests are shaped between dimer structures providing another scaffold enclosing detergents. Both scaffolds underline the potential of LL-37 to form defined peptide-lipid complexes in vivo. After adopting the activated peptide conformation LL-37 can polymerize and selectively extract bacterial lipids whereby the membrane is destabilized. The supramolecular fibril-like architectures formed in crystals can be reproduced in a peptide-lipid system after nanogold-labelled LL-37 interacted with lipid vesicles as followed by electron microscopy. We suggest that these supramolecular structures represent the LL-37-membrane active state. Collectively, our study provides new insights into the fascinating plasticity of LL-37 demonstrated at atomic resolution and opens the venue for LL-37-based molecules as novel antibiotics.

The Human LL-37(17-29) Antimicrobial Peptide Reveals a Functional Supramolecular Nanostructure

“…In contrast, structures of full-length LL-37, co-crystallized alone or with different lipids, resulted in different levels of assembly, including monomeric, dimeric, tetramers and fiber-like structure of oligomers (38). When co-crystallized with dodecylphosphocholine (PDB ID 5NNT) (38), full-length LL-37 showed a repetitive architecture of juxtaposed 'head-totail' amphipathic helices, with interactions mediated by detergent molecules. This structure formed a much looser packing compared to the LL-3717-29 structure, with only 45% of the helix buried within the protein assembly.…”

“…For comparison, in the PSMα3 cross-α structure, each helix is 62% buried in the fibril (19). In contrast, structures of full-length LL-37, co-crystallized alone or with different lipids, resulted in different levels of assembly, including monomeric, dimeric, tetramers and fiber-like structure of oligomers (38). When co-crystallized with dodecylphosphocholine (PDB ID 5NNT) (38), full-length LL-37 showed a repetitive architecture of juxtaposed 'head-totail' amphipathic helices, with interactions mediated by detergent molecules.…”

Self-assembly of glycerol monooleate with the antimicrobial peptide LL-37: a molecular dynamics study

“…17 Separation of the hydrophobic and hydrophilic groups into two different regions forces LL-37 to either oligomerize with other LL-37 peptides, a process that has recently been proposed as an initial step to destabilize membranes or to interact with the hydrophobic aliphatic chains of lipid membranes. 18,19 Once the concentration of the peptide on the microbe membranes reaches a threshold level, it starts to destabilize it, eventually leading to the pathogen's death. [19][20][21] Pathogens have evolved defense mechanisms against antimicrobial peptides, including the production of proteinases that can cleave LL-37 and either inhibit his activity or severely hamper its action.…”

“…18,19 Once the concentration of the peptide on the microbe membranes reaches a threshold level, it starts to destabilize it, eventually leading to the pathogen's death. [19][20][21] Pathogens have evolved defense mechanisms against antimicrobial peptides, including the production of proteinases that can cleave LL-37 and either inhibit his activity or severely hamper its action. 22 Sieprawska-Lupa, for instance, showed that Staphylococcus auresus's aureolysin (a metalloproteinase) was able to cleave and inactivate LL-37 and that Staphylococcus aureus strains with higher expression of metalloprotease were less susceptible to LL-37 activity.…”

Drug delivery systems designed to overcome antimicrobial resistance

“…In structural analysis, AMPs possess different secondary structures, often stabilized by disulfide bridges . They can be divided into four subgroups: α‐helical AMPs (eg, magainin and cathelicidin [LL‐37]), β‐sheet AMPs (eg, defensin, protegrin, tachyplesin, arenicin, extended AMPs (eg, indolicidin and bactenecin) (Figure ) . However, there are also other ways to classify AMPs as found in the AMP Databases …”