Self-assembly of spin-labeled antimicrobial peptides magainin 2 and PGLa in lipid bilayers.

The cationic antimicrobial peptides PGLa and magainin 2 (Mag2) are known for their antimicrobial activity and synergistic enhancement in antimicrobial and membrane leakage assays. Further use of peptides in combinatory therapy requires knowledge of the mechanisms of action of both individual peptides and their mixtures. Here, electron paramagnetic resonance (EPR), double electron-electron resonance (DEER, also known as PELDOR) and electron spin echo envelope modulation (ESEEM) spectroscopies were applied to study self-assembly and localization of spin-labeled PGLa and Mag2 in POPE/POPG membranes with a wide range of peptide/lipid ratios (P/L) from ∼1/1500 to 1/50. EPR and DEER data showed that both peptides tend to organize in clusters, which occurs already at the lowest peptide/lipid molar ratio of 1/1500 (0.067 mol%). For individual peptides, these clusters are quite dense with intermolecular distances of the order of ∼2 nm. In the presence of a synergistic peptide partner, these homo-clusters are transformed into lipid-diluted hetero-clusters. These clusters are characterized by a local surface density that is several times higher than expected from a random distribution. ESEEM data indicate a slightly different insertion depth of peptides in hetero-clusters when compared to homo-clusters.

Peptide Flexibility and the Hydrophobic Moment are Determinants to Evaluate the Clinical Potential of Magainins.

Using a flexibility prediction algorithm and in silico structural modeling, we have calculated the intrinsic flexibility of several magainin derivatives. In the case of magainin-2 (Mag-2) and magainin H2 (MAG-H2) we have found that MAG-2 is more flexible than its hydrophobic analog, Mag-H2. This affects the degree of bending of both peptides, with a kink around two central residues (R10, R11), whereas, in Mag-H2, W10 stiffens the peptide. Moreover, this increases the hydrophobic moment of Mag-H2, which could explain its propensity to form pores in POPC model membranes, which exhibit near-to-zero spontaneous curvatures. Likewise, the protective effect described in DOPC membranes for this peptide regarding its facilitation in pore formation would be related to the propensity of this lipid to form membranes with negative spontaneous curvature. The flexibility of another magainin analog (MSI-78) is even greater than that of Mag-2. This facilitates the peptide to present a kind of hinge around the central F12 as well as a C-terminal end prone to be disordered. Such characteristics are key to understanding the broad-spectrum antimicrobial actions exhibited by this peptide. These data reinforce the hypothesis on the determinant role of spontaneous membrane curvature, intrinsic peptide flexibility, and specific hydrophobic moment in assessing the bioactivity of membrane-active antimicrobial peptides.

Leaflet by Leaflet Synergistic Effects of Antimicrobial Peptides on Bacterial and Mammalian Membrane Models.

Antimicrobial peptides (AMPs) offer significant hope in the fight against antibiotic resistance. Operating via a mechanism different from that of antibiotics, they target the microbial membrane and ideally should damage it without impacting mammalian cells. Here, the interactions of two AMPs, magainin 2 and PGLa, and their synergistic effects on bacterial and mammalian membrane models were studied using electrochemical impedance spectroscopy, atomic force microscopy (AFM), and fluorescence correlation spectroscopy. Toroidal pore formation was observed by AFM when the two AMPs were combined, while individually AMP effects were confined to the exterior leaflet of the bacterial membrane analogue. Using microcavity-supported lipid bilayers, the diffusivity of each bilayer leaflet could be studied independently, and we observed that combined, the AMPs penetrate both leaflets of the bacterial model but individually each peptide had a limited impact on the proximal leaflet of the bacterial model. The impact of AMPs on a ternary, mammalian mimetic membrane was much weaker.

Antimicrobial peptide magainin 2-induced rupture of single giant unilamellar vesicles comprising E. coli polar lipids.

Most antimicrobial peptides (AMPs) damage the cell membrane of bacterial cells and induce rapid leakage of the internal cell contents, which is a main cause of their bactericidal activity. One of the AMPs, magainin 2 (Mag), forms nanopores in giant unilamellar vesicles (GUVs) comprising phosphatidylcholine (PC) and phosphatidylglycerol (PG), inducing leakage of fluorescent probes. In this study, to elucidate the Mag-induced pore formation in lipid bilayer region in E. coli cell membrane, we examined the interaction of Mag with single GUVs comprising E. coli polar lipids (E. coli-lipid-GUVs). First, we investigated the Mag-induced leakage of a fluorescent probe AF488 from single E. coli-lipid-GUVs, and found that Mag caused rupture of GUVs, inducing rapid AF488 leakage. The rate constant of Mag-induced GUV rupture increased with the Mag concentration. Using fluorescence microscopy with a time resolution of 5 ms, we revealed the GUV rupture process: first, a small micropore was observed in the GUV membrane, then the pore radius increased within 50 ms without changing the GUV diameter, the thickness of the membrane at the pore rim concomitantly increased, and eventually membrane aggregates were formed. Mag bound to only the outer monolayer of the GUV before GUV rupture, which increased the area of the GUV bilayer. We also examined the physical properties of E. coli-lipid-GUVs themselves. We found that the rate constant of the constant tension-induced rupture of E. coli-lipid-GUVs was higher than that of PG/PC-GUVs. Based on these results, we discussed the Mag-induced rupture of E. coli-lipid-GUVs and its mechanism.

Effect of lipid saturation on the topology and oligomeric state of helical membrane polypeptides.

Natural liquid crystalline membranes are made up of many different lipids carrying a mixture of saturated and unsaturated fatty acyl chains. Whereas in the past considerable attention has been paid to cholesterol content, the phospholipid head groups and the membrane surface charge the detailed fatty acyl composition was often considered less important. However, recent investigations indicate that the detailed fatty acyl chain composition has pronounced effects on the oligomerization of the transmembrane helical anchoring domains of the MHC II receptor or the membrane alignment of the cationic antimicrobial peptide PGLa. In contrast the antimicrobial peptides magainin 2 and alamethicin are less susceptible to lipid saturation. Using histidine-rich LAH4 designer peptides the high energetic contributions of lipid saturation in stabilizing transmembrane helical alignments are quantitatively evaluated. These observations can have important implications for the biological regulation of membrane proteins and should be taken into considerations during biophysical or structural experiments.

Single-Cell Analysis of the Antimicrobial and Bactericidal Activities of the Antimicrobial Peptide Magainin 2.

Antimicrobial peptides (AMPs) inhibit the proliferation of or kill bacterial cells. To measure these activities, several methods have been used, which provide only the average value of many cells. Here, we report the development of a method to examine the antimicrobial and bactericidal activities of AMPs at the single-cell level (i.e., single-cell analysis) and apply this strategy to examine the interaction of an AMP, magainin 2 (Mag), with Escherichia coli cells. Using this method, we monitored the proliferation of single cells on agar in a microchamber and measured the distribution of the number of cells in each microcolony using optical microscopy. For method A, we incubated cells in the presence of various concentrations of AMPs for 3 h. The fraction of microcolonies containing only a single cell, Psingle, increased with the Mag concentration and reached 1 at a specific concentration, which corresponded to the MIC. For method B, after the interaction of a cell suspension with an AMP for a specific time, an aliquot was diluted to stop the interaction, and the proliferation of single cells then was monitored after a 3-h incubation; this method permits the definition of Psingle(t), the fraction of dead cells after the interaction. For the interaction of Mag with E. coli cells, Psingle(t) increased with the interaction time, reaching ~1 at 10 and 20 min for 25 and 13 µM Mag, respectively. Thus, these results indicate that a short interaction time between Mag and E. coli cells is sufficient to induce bacterial cell death. IMPORTANCE To elucidate the activity of antimicrobial peptides (AMPs) against bacterial cells, it is important to estimate the interaction time that is sufficient to induce cell death. We have developed a method to examine the antimicrobial and bactericidal activities of AMPs at the single-cell level (i.e., single-cell analysis). Using this method, we monitored the proliferation of single cells on agar in a microchamber and measured the distribution of the number of cells in each microcolony using optical microscopy. We found that during the interaction of magainin 2 (Mag) with E. coli cells, the fraction of dead cells, Psingle(t), increased with the interaction time, rapidly reaching 1 (e.g., 10 min for 25 µM Mag). This result indicates that Mag induces cell death after a short time of interaction.

Concerted Rolling and Penetration of Peptides during Membrane Binding.

Peptide binding to membranes is common and fundamental in biochemistry and biophysics and critical for applications ranging from drug delivery to the treatment of bacterial infections. However, it is largely unclear, from a theoretical point of view, what peptides of different sequences and structures share in the membrane-binding and insertion process. In this work, we analyze three prototypical membrane-binding peptides (α-helical magainin, PGLa, and ß-hairpin tachyplesin) during membrane binding, using molecular details provided by Markov state modeling and microsecond-long molecular dynamics simulations. By leveraging both geometric and data-driven collective variables that capture the essential physics of the amphiphilic and cationic peptide-membrane interactions, we reveal how the slowest kinetic process of membrane binding is the dynamic rolling of the peptide from an attached to a fully bound state. These results not only add fundamental knowledge of the theory of how peptides bind to biological membranes but also open new avenues to study general peptides in more complex environments for further applications.

Anurans against SARS-CoV-2: A review of the potential antiviral action of anurans cutaneous peptides.

At the end of 2019, in China, clinical signs and symptoms of unknown etiology have been reported in several patients whose sample sequencing revealed pneumonia caused by the SARS-CoV-2 virus. COVID-19 is a disease triggered by this virus, and in 2020, the World Health Organization declared it a pandemic. Since then, efforts have been made to find effective therapeutic agents against this disease. Identifying novel natural antiviral drugs can be an alternative to treatment. For this reason, antimicrobial peptides secreted by anurans' skin have gained attention for showing a promissory antiviral effect. Hence, this review aimed to elucidate how and which peptides secreted by anurans' skin can be considered therapeutic agents to treat or prevent human viral infectious diseases. Through a literature review, we attempted to identify potential antiviral frogs' peptides to combat COVID-19. As a result, the Magainin-1 and -2 peptides, from the Magainin family, the Dermaseptin-S9, from the Dermaseptin family, and Caerin 1.6 and 1.10, from the Caerin family, are molecules that already showed antiviral effects against SARS-CoV-2 in silico. In addition to these peptides, this review suggests that future studies should use other families that already have antiviral action against other viruses, such as Brevinins, Maculatins, Esculentins, Temporins, and Urumins. To apply these peptides as therapeutic agents, experimental studies with peptides already tested in silico and new studies with other families not tested yet should be considered.

Magainin 2 and PGLa in bacterial membrane mimics III: Membrane fusion and disruption.

We previously speculated that the synergistically enhanced antimicrobial activity of Magainin 2 and PGLa is related to membrane adhesion, fusion, and further membrane remodeling. Here we combined computer simulations with time-resolved in vitro fluorescence microscopy, cryoelectron microscopy, and small-angle X-ray scattering to interrogate such morphological and topological changes of vesicles at nanoscopic and microscopic length scales in real time. Coarse-grained simulations revealed formation of an elongated and bent fusion zone between vesicles in the presence of equimolar peptide mixtures. Vesicle adhesion and fusion were observed to occur within a few seconds by cryoelectron microscopy and corroborated by small-angle X-ray scattering measurements. The latter experiments indicated continued and time-extended structural remodeling for individual peptides or chemically linked peptide heterodimers but with different kinetics. Fluorescence microscopy further captured peptide-dependent adhesion, fusion, and occasional bursting of giant unilamellar vesicles a few seconds after peptide addition. The synergistic interactions between the peptides shorten the time response of vesicles and enhance membrane fusogenic and disruption properties of the equimolar mixture compared with the individual peptides.

The effects of magainin 2-derived and rationally designed antimicrobial peptides on Mycoplasma pneumoniae.

Combating the spread of antimicrobial resistance (AMR) among bacteria requires a new class of antimicrobials, which desirably have a narrow spectrum because of their low propensity for the spread of AMR. Antimicrobial peptides (AMPs), which target the bacterial cell membrane, are promising seeds for novel antimicrobials because the cell membrane is essential for all cells. Previously, we reported the antimicrobial and haemolytic effects of a natural AMP, magainin 2 (Mag2), isolated from the skin of Xenopus laevis (the African clawed frog), four types of synthesised Mag2 derivatives, and three types of rationally designed AMPs on gram-positive and gram-negative bacteria. To identify novel antimicrobial seeds, we evaluated the effect of AMPs on Mycoplasma pneumoniae, which also exhibits AMR. We also evaluated the antimicrobial effects of an AMP, NK2A, which has been reported to have antimicrobial effects on Mycoplasma bovis, in addition to Mag2 and previously synthesised seven AMPs, on four strains of M. pneumoniae using colorimetric, biofilm, and killing assays. We found that three synthesised AMPs, namely 17base-Ac6c, 17base-Hybrid, and Block, had anti-M. pneumoniae (anti-Mp) effect at 8-30 µM, whereas others, including NK2A, did not have any such effect. For the further analysis, the membrane disruption activities of AMPs were measured by propidium iodide (PI) uptake assays, which suggested the direct interaction of AMPs to the cell membrane basically following the colorimetric, biofilm, and killing assay results. PI uptake assay, however, also showed the NK2A strong interaction to cell membrane, indicating unknown anti-Mp determinant factors related to the peptide sequences. Finally, we conclude that anti-Mp effect was not simply determined by the membrane disruption activities of AMPs, but also that the sequence of AMPs were important for killing of M. pneumoniae. These findings would be helpful for the development of AMPs for M. pneumoniae.

Lipid tails modulate antimicrobial peptide membrane incorporation and activity.

Membrane disrupting antimicrobial peptides (AMPs) are often amphipathic peptides that interact directly with lipid bilayers. AMPs are generally thought to interact mostly with lipid head groups, but it is less clear how the lipid alkyl chain length and saturation modulate interactions with membranes. Here, we used native mass spectrometry to measure the stoichiometry of three different AMPs-LL-37, indolicidin, and magainin-2-in lipid nanodiscs. We also measured the activity of these AMPs in unilamellar vesicle leakage assays. We found that LL-37 formed specific hexamer complexes but with different intermediates and affinities that depended on the bilayer thickness. LL-37 was also most active in lipid bilayers containing longer, unsaturated lipids. In contrast, indolicidin incorporated to a higher degree into more fluid lipid bilayers but was more active with bilayers with thinner, less fluid lipids. Finally, magainin-2 incorporated to a higher degree into bilayers with longer, unsaturated alkyl chains and showed more activity in these same conditions. Together, these data show that higher amounts of peptide incorporation generally led to higher activity and that AMPs tend to incorporate more into longer unsaturated lipid bilayers. However, the activity of AMPs was not always directly related to amount of peptide incorporated.

Lipid saturation and head group composition have a pronounced influence on the membrane insertion equilibrium of amphipathic helical polypeptides.

The histidine-rich peptides of the LAH4 family were designed using cationic antimicrobial peptides such as magainin and PGLa as templates. The LAH4 amphipathic helical sequences exhibit a multitude of interesting biological properties such as antimicrobial activity, cell penetration of a large variety of cargo and lentiviral transduction enhancement. The parent peptide associates with lipid bilayers where it changes from an orientation along the membrane interface into a transmembrane configuration in a pH-dependent manner. Here we show that LAH4 adopts a transmembrane configuration in fully saturated DMPC membranes already at pH 3.5, i.e. much below the pKa of the histidines whereas the transition pH in POPC correlates closely with histidine neutralization. In contrast in POPG membranes the in-planar configuration is stabilized by about one pH unit. The differences in pH can be converted into energetic contributions for the in-plane to transmembrane transition equilibrium, where the shift in the transition pH due to lipid saturation corresponds to energies which are otherwise obtained by the exchange of several cationic with hydrophobic residues. A similar dependence on lipid saturation has also been observed when the PGLa and magainin antimicrobial peptides interact within lipid bilayers suggesting that the quantitative evaluation presented in this paper also applies to other membrane polypeptides.

Investigation of the Action of Peptides on Lipid Membranes.

Calorimetric and incoherent neutron scattering methods were employed to investigate the action of magainin 2 and PGLa peptides on the phase behavior and molecular dynamics of lipids mimicking cytoplasmic membranes of Gram-negative bacteria. The impact of the peptides, tested individually and cooperatively by differential scanning calorimetry, presented a broadened peak, sometimes with a second shoulder, depicting the phase transition temperature around 21 °C. Neutron scattering revealed a small but significant variation of the membrane dynamics due to the peptides in both in-plane and out-of-plane directions. Although we did not find a clear hint for synergy in the interplay of the two peptides, the calorimetric and neutron data give compatible results in terms of a decrease of the enthalpy due to the presence of the peptides, which destabilize the membrane. The dynamics in the two directions was differentiated when the individual peptides were added to the membranes, but the impact was smaller when both peptides were added together.

A magainin-2 like bacteriocin BpSl14 with anticancer action from fish gut Bacillus safensis SDG14.

Bacteriocins are gaining utmost importance in antimicrobial and chemotherapy due to their diverse structure and activity. This study centres on magainin-2 like bacteriocin with anticancer action, produced by Bacillus safensis strain SDG14 isolated from gut of marine fish Sardinella longiceps. The purified bacteriocin designated as BpSl14 was thermostable and pH tolerant. The molecular weight of BpS114 was estimated to be 6061.2 Da using MALDI-ToF MS. The partial primary sequence was elucidated by peptide mass fingerprinting using MALDI MS/MS. The tertiary structure of the partial sequence was similar to that of two magainin-2 α-helices joined together by extended indolicidin. The BpSl14 protein inhibited the cells of lung carcinoma, one of the deadliest cancers. Docking studies conducted with DR5 and TGF-ß, two of the most prominent apoptotic receptors in adenocarcinoma, also proved the anti-apoptotic action of BpSl14.

Ligand-decoration determines the translational and rotational dynamics of nanoparticles on a lipid bilayer membrane.

Nanoparticles (NPs) promise a huge potential for clinical diagnostic and therapeutic applications. However, nano-bio (e.g., the NP-cell membrane) interactions and underlying mechanisms are still largely elusive. In this study, two types of congeneric peptides, namely PGLa and magainin 2 (MAG2), with similar membrane activities were employed as model ligands for NP decoration, and the diffusion behaviours (including both translation and rotation) of the ligand-decorated NPs on a lipid bilayer membrane were studied via molecular dynamics simulations. It was found that, although both PGLa- and MAG2-coated NPs showed alternatively "hopping" and "jiggling" diffusions, the PGLa-coated ones had an enhanced circling at the hopping stage, while a much confined circling at the jiggling stage. In contrast, the MAG2-coated NPs demonstrated constant circling tendencies throughout the diffusion process. Such differences in the coupling between translational and rotational dynamics of these two types of NPs are ascribed to the different ligand-lipid interactions of PGLa and MAG2, in which the PGLa ligands prefer to vertically insert into the membrane, while MAG2 tends to lie flat on the membrane surface. Our results are helpful for the understanding the underlying associations between the NP motions and their interfacial membrane interactions, and shed light on the possibility of regulating NP behaviours on a cellular surface for better biomedical uses.

Development of Antimicrobial Stapled Peptides Based on Magainin 2 Sequence.

Magainin 2 (Mag2), which was isolated from the skin of the African clawed frog, is a representative antimicrobial peptide (AMP) that exerts antimicrobial activity via microbial membrane disruption. It has been reported that the helicity and amphipathicity of Mag2 play important roles in its antimicrobial activity. We investigated and recently reported that 17 amino acid residues of Mag2 are required for its antimicrobial activity, and accordingly developed antimicrobial foldamers containing α,α-disubstituted amino acid residues. In this study, we further designed and synthesized a set of Mag2 derivatives bearing the hydrocarbon stapling side chain for helix stabilization. The preferred secondary structures, antimicrobial activities, and cell-membrane disruption activities of the synthesized peptides were evaluated. Our analyses revealed that hydrocarbon stapling strongly stabilized the helical structure of the peptides and enhanced their antimicrobial activity. Moreover, peptide 2 stapling between the first and fifth position from the N-terminus showed higher antimicrobial activity than that of Mag2 against both gram-positive and gram-negative bacteria without exerting significant hemolytic activity. To investigate the modes of action of tested peptides 2 and 8 in antimicrobial and hemolytic activity, electrophysiological measurements were performed.

Palmitoylation of Membrane-Penetrating Magainin Derivatives Reinforces Necroptosis in A549 Cells Dependent on Peptide Conformational Propensities.

Anticancer lipopeptides (ACLPs) are considered promising alternatives to combat resistant cancer cells, but the influence of peptide conformational propensity alone on their selectivity and mechanism remains obscure. In this study, we developed N-palmitoylated MK5E (P1MK5E) and MEK5 (P1MEK5) that have the same composition of 23 residues undergoing the pH-dependent structural alterations but differ in the conformational tendency of their amino acid composites. Nonlipidated peptides were readily accumulated in the A549 cell nucleus by the direct membrane translocation and the heparan sulfate-mediated endocytosis than the lipid-raft-dependent pathway. The increased hydrophobicity favored the amino acid-position-dependent folding of P1MK5E and P1MEK5, respectively, toward the α-helical coiled-coil nanofibrils and amyloidlike ß-protofibrils. At the close concentrations (∼7.5 µM) to the toxic effects of doxorubicin (DOX), P1MK5E exhibited (i) an increased anticancer toxicity through a time-dependent S-phase arrest, (ii) enhanced plasma membrane permeability, and (iii) dose-dependent changes in the cell death characteristic features in the A549 cells relative to P1MEK5 that was almost inactive at ∼75 µM. These observations were in accordance with the TNF-α-mediated necroptotic signaling in the c-MYC/PARP1-overexpressed A549 cells exposed to P1MK5E and accompanied by the ultrastructure of plasma membrane protrusions, extensive endoplasmic reticulum (ER) membrane expansion, mitochondrial swelling, and the formation of distinct cytoplasmic vacuolation. The structural results and the bioactivity behaviors, herein, declared the significance of α-helical propensity in the peptide sequence and the nanostructure morphologies of self-assembling ACLPs upon the selectivity and enhanced anticancer effectiveness, which notably holds promise in the design and development of efficient therapeutics for cancer.

Review: Examining the Natural Role of Amphibian Antimicrobial Peptide Magainin.

Host defense peptides (HDPs) are a group of antimicrobial peptides (AMPs) that are crucial components of the innate immune system of many different organisms. These small peptides actively kill microbes and prevent infection. Despite the presence of AMPs in the amphibian immune system, populations of these organisms are in decline globally. Magainin is an AMP derived from the African clawed frog (Xenopus laevis) and has displayed potent antimicrobial effects against a wide variety of microbes. Included in this group of microbes are known pathogens of the African clawed frog and other amphibian species. Arguably, the most deleterious amphibious pathogen is Batrachochytrium dendrobatidis, a chytrid fungus. Investigating the mechanism of action of magainin can help understand how to effectively fight off infection. By understanding amphibian AMPs' role in the frog, a potential conservation strategy can be developed for other species of amphibians that are susceptible to infections, such as the North American green frog (Rana clamitans). Considering that population declines of these organisms are occurring globally, this effort is crucial to protect not only these organisms but the ecosystems they inhabit as well.

Highly synergistic antimicrobial activity of magainin 2 and PGLa peptides is rooted in the formation of supramolecular complexes with lipids.

Magainin 2 and PGLa are cationic, amphipathic antimicrobial peptides which when added as equimolar mixture exhibit a pronounced synergism in both their antibacterial and pore-forming activities. Here we show for the first time that the peptides assemble into defined supramolecular structures along the membrane interface. The resulting mesophases are quantitatively described by state-of-the art fluorescence self-quenching and correlation spectroscopies. Notably, the synergistic behavior of magainin 2 and PGLa correlates with the formation of hetero-domains and an order-of-magnitude increased membrane affinity of both peptides. Enhanced membrane association of the peptide mixture is only observed in the presence of phophatidylethanolamines but not of phosphatidylcholines, lipids that dominate bacterial and eukaryotic membranes, respectively. Thereby the increased membrane-affinity of the peptide mixtures not only explains their synergistic antimicrobial activity, but at the same time provides a new concept to increase the therapeutic window of combinatorial drugs.

Traceless native chemical ligation of lipid-modified peptide surfactants by mixed micelle formation.

Biology utilizes multiple strategies, including sequestration in lipid vesicles, to raise the rate and specificity of chemical reactions through increases in effective molarity of reactants. We show that micelle-assisted reaction can facilitate native chemical ligations (NCLs) between a peptide-thioester - in which the thioester leaving group contains a lipid-like alkyl chain - and a Cys-peptide modified by a lipid-like moiety. Hydrophobic lipid modification of each peptide segment promotes the formation of mixed micelles, bringing the reacting peptides into close proximity and increasing the reaction rate. The approach enables the rapid synthesis of polypeptides using low concentrations of reactants without the need for thiol catalysts. After NCL, the lipid moiety is removed to yield an unmodified ligation product. This micelle-based methodology facilitates the generation of natural peptides, like Magainin 2, and the derivatization of the protein Ubiquitin. Formation of mixed micelles from lipid-modified reactants shows promise for accelerating chemical reactions in a traceless manner.