Enhanced Cationic Charge is a Key Factor in Promoting Staphylocidal Activity of α-Melanocyte Stimulating Hormone via Selective Lipid Affinity

Abstract

The steady rise in antimicrobial resistance poses a severe threat to global public health by hindering treatment of an escalating spectrum of infections. We have previously established the potent activity of α-MSH, a 13 residue antimicrobial peptide, against the opportunistic pathogen Staphylococcus aureus. Here, we sought to determine whether an increase in cationic charge in α-MSH could contribute towards improving its staphylocidal potential by increasing its interaction with anionic bacterial membranes. For this we designed novel α-MSH analogues by replacing polar uncharged residues with lysine and alanine. Similar to α-MSH, the designed peptides preserved turn/random coil conformation in artificial bacterial mimic 1,2-dimyristoyl-sn-glycero-3-phosphocholine:1,2-dimyristoyl-sn-glycero-3-phospho-rac-(1-glycerol) (7:3, w/w) vesicles and showed preferential insertion in the hydrophobic core of anionic membranes. Increased cationic charge resulted in considerable augmentation of antibacterial potency against MSSA and MRSA. With ~18-fold better binding than α-MSH to bacterial mimic vesicles, the most charged peptide KKK-MSH showed enhanced membrane permeabilization and depolarization activity against intact S. aureus. Scanning electron microscopy confirmed a membrane disruptive mode of action for KKK-MSH. Overall, increasing the cationic charge improved the staphylocidal activity of α-MSH without compromising its cell selectivity. The present study would help in designing more effective α-MSH-based peptides to combat clinically relevant staphylococcal infections.

Unifying structural signature of eukaryotic α-helical host defense peptides

“…ability of a peptide to induce NGC in microbial target membranes, such that deviations away from the signature diminishes their ability to do so. Positive charge is thought to contribute to antimicrobial selectivity, given relative electronegative charge and inward-rectified potential (δψ) of prokaryotic cell mem-branes or mitochondria of eukaryotic pathogens (2,(25)(26)(27)(28). Moreover, the high concentration of lipids with negative intrinsic curvature in a target membrane, as is the case for most bacteria, also contributes to specific activity of αHDPs (29-31) by modifying the Gaussian modulus.…”

Novel Miniature Membrane Active Lipopeptidomimetics against Planktonic and Biofilm Embedded Methicillin-Resistant Staphylococcus aureus

“…For the active peptidomimetics cytotoxicity was measured against 3T3 murine fibroblast cell line using a previously defined MTT assay with slight modifications 26 . Briefly, the cells were grown to over 75% confluence in DMEM supplemented with 10% FBS and antibiotics (1 × Anti-anti) at 37 °C in 5% CO 2 atmosphere.…”

“…Previously, in our efforts to design cost-effective (short), resistance proof and membrane disruptive antibacterial peptides/peptidomimetics, we have explored the effect of increasing cationic charge by rational substitution of amino acids or modulation of hydrophobicity by using various N- and C-terminal unnatural modifications 23 – 26 . In our present work, we sought to develop short (5-mer), membrane disruptive, protease-stable peptidomimetics by incorporating unnatural moieties in the middle of the sequences to better fine tune hydrophobicity and thus activity/selectivity.…”

N-myristoylation of Antimicrobial Peptide CM4 Enhances Its Anticancer Activity by Interacting With Cell Membrane and Targeting Mitochondria in Breast Cancer Cells

“…Erythrocytes were isolated from fresh mouse blood cells by centrifugation at 1000 g for 10 minutes (min) and washed three times with PBS. Hemolytic activity was evaluated to the method described previously ( Singh et al, 2016 ). Briefly, erythrocytes (final concentration 4% v/v) were treated with myristoyl-CM4 or CM4 for 1 h at 37°C, followed by centrifugation at 1000 g for 5 min.…”