Goldilocks Dilemma: LPS Works Both as the Initial Target and a Barrier for the Antimicrobial Action of Cationic AMPs on E. coli

Antimicrobial peptides (AMPs) are generally membrane-active compounds that physically disrupt bacterial membranes. Despite extensive research, the precise mode of action of AMPs is still a topic of great debate. This work demonstrates that the initial interaction between the Gram-negative E. coli and AMPs is driven by lipopolysaccharides (LPS) that act as kinetic barriers for the binding of AMPs to the bacterial membrane. A combination of SPR and NMR experiments provide evidence suggesting that cationic AMPs first bind to the negatively charged LPS before reaching a binding place in the lipid bilayer. In the event that the initial LPS-binding is too strong (corresponding to a low dissociation rate), the cationic AMPs cannot effectively get from the LPS to the membrane, and their antimicrobial potency will thus be diminished. On the other hand, the AMPs must also be able to effectively interact with the membrane to exert its activity. The ability of the studied cyclic hexapeptides to bind LPS and to translocate into a lipid membrane is related to the nature of the cationic charge (arginine vs. lysine) and to the distribution of hydrophobicity along the molecule (alternating vs. clumped tryptophan)

Label-free measurement of antimicrobial peptide interactions with lipid vesicles and nanodiscs using microscale thermophoresis

One strategy to combat antimicrobial resistance is the discovery of new classes of antibiotics. Most antibiotics will at some point interact with the bacterial membrane to either interfere with its integrity or to cross it. Reliable and efficient tools for determining the dissociation constant for membrane binding (KD) and the partitioning coefficient between the aqueous- and membrane phases (KP) are therefore important tools for discovering and optimizing antimicrobial hits. Here we demonstrate that microscale thermophoresis (MST) can be used for label-free measurement of KD by utilising the intrinsic fluorescence of tryptophan and thereby removing the need for chromophore labelling. As proof of principle, we have used the method to measure the binding of a set of small cyclic AMPs to large unilamellar vesicles (LUVs) and two types of lipid nanodiscs assembled by styrene maleic acid (SMA) and quaternary ammonium SMA (SMA-QA). The measured KD values correlate well with the corresponding measurements using surface plasmon resonance (SPR), also broadly reflecting the tested AMPs’ minimal inhibition concentration (MIC) towards S. aureus and E. coli. We conclude that MST is a promising method for fast and cost-efficient detection of peptide-lipid interactions or mapping of sample conditions in preparation for more advanced studies that rely on expensive sample preparation, labelling and/or instrument time

Anti-proliferative activity of a novel tricyclic triterpenoid acid from Commiphora africana resin against four human cancer cell lines

Myrrh, a resin derived from the damaged bark of Commiphora genus, has traditionally been used for treatment of various human diseases, such as amenorrhea, ache, tumors, fever, and stomach pains. In spite of this widespread use of the myrrh in Ethiopia, the pharmacological activity and chemical composition have not been studied in detail. A new tricyclic triterpene acid (3S,4S,14S,7E,17E,21Z)-3,30-dihydroxypodioda-7,17,21-trien-4-carboxylic acid (commafric A) has been isolated from a crude methanolic extract of Commiphora africana (A. Rich.) Engl. resin along with the known pentacyclic triterpene α-amyrin. The structure of commafric A was characterized using diferent spectroscopic techniques such as 1D and 2D NMR, IR, and VCD combined with computations. The anti-proliferative activity of both isolated compounds was evaluated using SRB based colorimetric cellular assay against four human cancer cell lines. Etoposide was used as a positive control. Commafric A showed signifcant anti-proliferative efects against non-small cell lung cancer (A549) with IC50 values of 4.52 μg/ml. The pentacyclic triterpene α-amyrin showed a weak antiproliferative activity against A2780 (ovarian cancer), MIA-PaCa-2 (pancreatic cancer), and SNU638 (stomach cancer) cell lines tested with IC50 values ranging 9.28 to 28.22 μg/ml. Commafric A possessed anti-proliferative activity against non-small cell lung cancer (A549), which suggests that commafric A has potential to be further optimized being a lead compound in the search for new drugs against cancer diseases