Electrochemiluminescence Imaging of Liposome Permeabilization by an Antimicrobial Peptide: Melittin

Abstract

The permeabilization of liposomes by melittin, an antimicrobial peptide (AMP), has been studied by an electrochemiluminescence (ECL) imaging strategy. The methodology consisted first of encapsulating ECL reagents in sealed giant asymmetrical liposomes (100 μm in diameter) made of DOPG/DOPC phospholipids (i.e., 1,2-dioleoyl-sn-glycerol-3-phospho-(1′-rac-glycerol) sodium salt/1,2-dioleolyl-sn-glycero-3-phosphocholine). Then liposomes were placed on an indium tin oxide electrode coated with poly-l-lysine to avoid any membrane poration/permeabilization through polarization of the electrode surface. Finally, the addition of melittin (from 10 μM to 100 nM in concentration) enabled the permeabilization of the lipid membrane followed by the liposome content release and subsequent light generation through the ECL reagents oxidation processes. Interestingly, at a melittin concentration of 10 μM, two successive leakages occurring on the same liposome could be imaged. Combination of ECL and photoluminescence imaging allowed comprehensive monitoring of the permeabilization and content release of a single liposome. This ECL imaging approach opens interesting perspectives to characterize the instant release of vesicle content upon permeabilization by AMPs or other membrane-active species.

The current landscape of the antimicrobial peptide melittin and its therapeutic potential

“…Melittin can be applied to monitor lipid-protein interactions in membranes ( 5 , 14 ). Interestingly, electrochemiluminescence (ECL) imaging efficiently characterize the instant release of vesicle content via melittin-triggered permeabilization ( 15 ). Melittin is also applied for the studies related to calmodulin (CaM), a sensor protein of calcium ions, which is regarded as hub protein to bind and modulate more than 200 target proteins in a concentration-dependent manner of calcium ions.…”

Wireless rotating bipolar electrochemiluminescence for enzymatic detection

“…[9][10][11][12][13][14] Thus, ECL generates an optical readout opening various bioanalytical and microscopy opportunities. [15][16][17][18][19][20][21] In the analytical applications of ECL, two main strategies were employed in the open bipolar configurations: (i) the analytical target is involved directly or indirectly in the ECL mechanism and the ECL intensity generated at one pole of the BE is proportional to its concentration; (ii) the redox reaction of the analyte occurs at a given pole named sensing pole and is coupled with the counter redox reaction generating the ECL signal at the opposite pole named reporting pole. 11,22 This latter strategy was introduced by Crooks and co-workers.…”

Shadow electrochemiluminescence imaging of giant liposomes opening at polarized electrodes

“…13 In another relevant example, a dual functional electroactive and fluorescent probe was used for coupled measurements of vesicular exocytosis with high spatial and temporal resolution. 14 Recently, we developed an original approach based on electrogenerated chemiluminescence (also called electrochemiluminescence, ECL) 15,16 allowing the imaging of liposome permeabilization processes triggered either by electroporation 17 or by the presence of the melittin AMP, 18 an anti-microbial peptide. On the one hand, liposomes (i.e., vesicles composed of a phospholipid bilayer) are widely investigated due to their increasing interest not only in active and passive lipid barrier crossings, but also in fields as diverse as targeted drug delivery, [19][20][21] bioreactors, [22][23][24] membrane protein science, [25][26][27] and artificial cells.…”