Peptides In Bilayers Research Articles

Examination of the Interaction between a Membrane Active Peptideand Artificial Bilayers by Dual Polarisation Interferometry.

Examining the interaction of peptides with lipid bilayers to determine binding kinetics is often performed using surface plasmon resonance (SPR). Here we describe the technique of dual polarisation interferometry (DPI) that provides not only information on the kinetics of the peptide binding to the bilayer, but also how the peptide affects the lipid order of the bilayer.

Computational Design of Membrane Curvature-Sensing Peptides.

Computer simulations have become an indispensable tool in studying molecular biological systems. The unmatched spatial and temporal resolution that it offers enables for microscopic-level views into the dynamics and mechanics of biological systems. Recent advances in hardware resources have also opened up to computer simulations the investigation of longer timescale biological processes and larger systems. The study of membrane proteins or peptides especially benefits from simulations due to difficulties related to crystallization of such proteins in a membrane environment. In this chapter, we outline the method of molecular dynamics and how it is applied to simulations that involve a peptide and lipid bilayers. In particular, the simulation of a membrane-curvature sensing peptide is examined, and ways of employing computational simulations to design such peptides are discussed.

Effect of lipid composition and amino acid sequence upon transmembrane peptide-accelerated lipid transleaflet diffusion (flip-flop)

We examined how hydrophobic peptide-accelerated transleaflet lipid movement (flip-flop) was affected by peptide sequence and vesicle composition and properties. A peptide with a completely hydrophobic sequence had little if any effect upon flip-flop. While peptides with a somewhat less hydrophobic sequence accelerated flip-flop, the half-time remained slow (hours) with substantial (0.5mol%) peptide in the membranes. It appears that peptide-accelerated lipid flip-flop involves a rare event that may reflect a rare state of the peptide or lipid bilayer. There was no simple relationship between peptide overall hydrophobicity and flip-flop. In addition, flip-flop was not closely linked to whether the peptides were in a transmembrane or non-transmembrane (interfacial) inserted state. Flip-flop was also not associated with peptide-induced pore formation. We found that peptide-accelerated flip-flop is initially faster in small (highly curved) unilamellar vesicles relative to that in large unilamellar vesicles. Peptide-accelerated flip-flop was also affected by lipid composition, being slowed in vesicles with thick bilayers or those containing 30% cholesterol. Interestingly, these factors also slow spontaneous lipid flip-flop in the absence of peptide. Combined with previous studies, the results are most consistent with acceleration of lipid flip-flop by peptide-induced thinning of bilayer width.

Role of the Cationic C-Terminal Segment of Melittin on Membrane Fragmentation.

The widespread distribution of cationic antimicrobial peptides capable of membrane fragmentation in nature underlines their importance to living organisms. In the present work, we determined the impact of the electrostatic interactions associated with the cationic C-terminal segment of melittin, a 26-amino acid peptide from bee venom (net charge +6), on its binding to model membranes and on the resulting fragmentation. In order to detail the role played by the C-terminal charges, we prepared a melittin analogue for which the four cationic amino acids in positions 21-24 were substituted with the polar residue citrulline, providing a peptide with the same length and amphiphilicity but with a lower net charge (+2). We compared the peptide bilayer affinity and the membrane fragmentation for bilayers prepared from 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC)/1,2-dipalmitoyl-sn-glycero-3-phospho-l-serine (DPPS) mixtures. It is shown that neutralization of the C-terminal considerably increased melittin affinity for zwitterionic membranes. The unfavorable contribution associated with transferring the cationic C-terminal in a less polar environment was reduced, leaving the hydrophobic interactions, which drive the peptide insertion in bilayers, with limited counterbalancing interactions. The presence of negatively charged lipids (DPPS) in bilayers increased melittin binding by introducing attractive electrostatic interactions, the augmentation being, as expected, greater for native melittin than for its citrullinated analogue. The membrane fragmentation power of the peptide was shown to be controlled by electrostatic interactions and could be modulated by the charge carried by both the membrane and the lytic peptide. The analysis of the lipid composition of the extracted fragments from DPPC/DPPS bilayers revealed no lipid specificity. It is proposed that extended phase separations are more susceptible to lead to the extraction of a lipid species in a specific manner than a specific lipid-peptide affinity. The present work on the lipid extraction by melittin and citrullinated melittin with model membranes emphasizes the complex relation between the affinity, the lipid extraction/membrane fragmentation, and the lipid specificity.

Design of Asymmetric Peptide Bilayer Membranes.

Energetic insights emerging from the structural characterization of peptide cross-β assemblies have enabled the design and construction of robust asymmetric bilayer peptide membranes. Two peptides differing only in their N-terminal residue, phosphotyrosine vs lysine, coassemble as stacks of antiparallel β-sheets with precisely patterned charged lattices stabilizing the bilayer leaflet interface. Either homogeneous or mixed leaflet composition is possible, and both create nanotubes with dense negative external and positive internal solvent exposed surfaces. Cross-seeding peptide solutions with a preassembled peptide nanotube seed leads to domains of different leaflet architecture within single nanotubes. Architectural control over these cross-β assemblies, both across the bilayer membrane and along the nanotube length, provides access to highly ordered asymmetric membranes for the further construction of functional mesoscale assemblies.

The Alzheimer's disease Aβ peptide binds to the anionic DMPS lipid bilayer

We have applied isobaric–isothermal replica exchange molecular dynamics (REMD) and the all-atom explicit water model to study binding of Aβ10–40 peptide to the anionic DMPS bilayer. To provide comparison with a zwitterionic bilayer, we used our previous REMD simulations probing binding of the same peptide to the DMPC bilayer. Using two sets of simulations, we comparatively analyzed the equilibrium Aβ conformational ensemble, peptide–bilayer interactions, and changes in the bilayer structure induced by Aβ binding. Our results are six-fold. (1) Binding to the DMPS bilayer triggers the formation of stable helix in the Aβ C-terminal, although the helix-inducing effect caused by DMPS lipids is weaker than that of DMPC. (2) Compared to the DMPC-bound Aβ monomer, the anionic bilayer weakens intrapeptide interactions, particularly, formed by charged amino acids. (3) Binding of Aβ peptide to the DMPS bilayer is primarily governed by electrostatic interactions between charged amino acids and charged lipid groups. In contrast, these interactions play minor role in Aβ binding to the DMPC bilayer. (4) Aβ peptide generally resides on the DMPS bilayer surface causing relatively minor bilayer thinning. The opposite scenario applies to Aβ binding to the DMPC bilayer. (5) In contrast to DMPC simulations, Aβ largely expels anionic lipids from its binding “footprint” forming a ring of charged amino acids mixed with charged lipid groups around the peptide. (6) Aβ binding disorders proximal DMPS lipids more strongly than their DMPC counterparts. Our simulations show that Aβ monomers fail to perturb anionic or zwitterionic bilayers across both leaflets.

Membrane interaction of antimicrobial peptides using E. coli lipid extract as model bacterial cell membranes and SFG spectroscopy

Supported lipid bilayers are used as a convenient model cell membrane system to study biologically important molecule–lipid interactions in situ. However, the lipid bilayer models are often simple and the acquired results with these models may not provide all pertinent information related to a real cell membrane. In this work, we use sum frequency generation (SFG) vibrational spectroscopy to study molecular-level interactions between the antimicrobial peptides (AMPs) MSI-594, ovispirin-1 G18, magainin 2 and a simple 1,2-dipalmitoyl-d62-sn-glycero-3-phosphoglycerol (dDPPG)/1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoglycerol (POPG) bilayer. We compared such interactions to those between the AMPs and a more complex dDPPG/Escherichia coli (E. coli) polar lipid extract bilayer. We show that to fully understand more complex aspects of peptide–bilayer interaction, such as interaction kinetics, a heterogeneous lipid composition is required, such as the E. coli polar lipid extract. The discrepancy in peptide–bilayer interaction is likely due in part to the difference in bilayer charge between the two systems since highly negative charged lipids can promote more favorable electrostatic interactions between the peptide and lipid bilayer. Results presented in this paper indicate that more complex model bilayers are needed to accurately analyze peptide–cell membrane interactions and demonstrates the importance of using an appropriate lipid composition to study AMP interaction properties.

Generation of Electrochemical Gradient from Peptide Self-Assembly

The nucleating core of the Amyloid β peptide associated with Alzheimer's disease is able to organize into a peptide bilayer maintaining dimensions similar to biological phospholipid membranes. However, the dynamics of the hydrogen-bonded peptides are certainly different from the flexible alkanes of the lipid membranes, and this crystallinity prompted us to consider their dynamics and functional potential. Here we will review these unique surfaces built on cross-β structures of amyloid, and extend that pattern to mixing of positively- and negatively-charged surfaces to give even higher order architectures with high electrochemical potential across the 4 nm membrane. Further, we have developed EFM analyses to map the charge distribution and explored novel energy or electron transduction reactions. Our results show that these peptide membrane scaffolds are dynamic, and their ability to self-organize offers a new opportunity for engineering specific molecular recognition elements into the peptide assemblies and well-ordered materials.

Monitoring of Single Vesicle Cytochrome-c Release Illuminates BAK as a Novel Target of Aβ Oligomers

Monitoring of Single Vesicle Cytochrome-c Release Illuminates BAK as a Novel Target of Aβ Oligomers

Lipid-like self-assembling peptide nanovesicles for drug delivery.

Amphiphilic self-assembling peptides are functional materials, which, depending on the amino acid sequence, the peptide length, and the physicochemical conditions, form a variety of nanostructures including nanovesicles, nanotubes, and nanovalves. We designed lipid-like peptides with an aspartic acid or lysine hydrophilic head and a hydrophobic tail composed of six alanines (i.e., ac-A6K-CONH2, KA6-CONH2, ac-A6D-COOH, and DA6-COOH). The resulting novel peptides have a length similar to biological lipids and form nanovesicles at physiological conditions. AFM microscopy and light scattering analyses of the positively charged lipid-like ac-A6K-CONH2, KA6-CONH2 peptide formulations showed individual nanovesicles. The negatively charged ac-A6D-COOH and DA6-COOH peptides self-assembled into nanovesicles that formed clusters that upon drying were organized into necklace-like formations of nanovesicles. Encapsulation of probe molecules and release studies through the peptide bilayer suggest that peptide nanovesicles may be good candidates for sustained release of pharmaceutically active hydrophilic and hydrophobic compounds. Lipid-like peptide nanovesicles represent a paradigm shifting system that may complement liposomes for the delivery of diagnostic and therapeutic agents.

The Structure of Cross‐β Tapes and Tubes Formed by an Octapeptide, αSβ1

Elaborate morphology: The αSβ1 peptide, a fragment of α-synuclein, assembles into flat tapes consisting of a peptide bilayer, which can be modeled based on the cross-β structure found in amyloid proteins. The tapes are stabilized by hydrogen bonding, whilst the amphiphilic nature of the peptide results in the thin bilayer structure. To further stabilize the structure, these tapes may twist to form helical tapes, which subsequently close into nanotubes.

Binding of β-Amyloid (1-42) Peptide to Negatively Charged Phospholipid Membranes in the Liquid-Ordered State

Plaques formed by aggregation of amyloid β peptide (Aβ 42) are characteristic of Alzheimer's disease. The mechanism of plaque formation from Aβ 42 monomers, oligomers and fibrils is not understood, although there are data indicating that negatively-charged membrane surfaces catalyze the process. It has also been suggested that the pathogenic peptides are originated in raft-like microdomains in the liquid-ordered state. In order to explore the initial stages of the process, we have studied the interaction of Aβ 42 in the monomeric form with lipid monolayers and bilayers in the liquid-ordered state, composed of equimolar brain sphingomyelin and cholesterol, to which either 5 or 20 mol% phosphatidic acid (PA) were added. Molecular dynamics simulations of the system were performed, as well as experimental measurements of changes in surface pressure at the air-water interface, and isothermal calorimetry of peptide-vesicle interactions. In the absence of the negatively charged PA interaction is weak and it cannot be detected calorimetrically. However in the presence of PA interaction is detected by all three methods and in all cases interaction is strongest with the lipid composition containing 5 mol% PA. Molecular dynamics pictures are different for the low- and highcharge bilayers, in the former case the peptide is bound through many contact points to the bilayer, while for the bilayer containing 20 mol% PA only a small fragment of the peptide appears to be bound. Interestingly, the calorimetric data indicate decreased entropy upon binding only for the system predicted to contain highly bound peptide. Moreover the molecular dynamics methods indicate the development of a β-sheet structure by the peptide in bilayers containing 5% PA, while in the presence of 20 mol% PA the peptide retains a partially helical conformation.

Membrane-Perturbing Activities of KL4-Related Surfactant Peptides

KL4 is a 21-residue peptide proposed as a potential substitute of pulmonary surfactant protein SP-B in synthetic surfactants, intended for the treatment of respiratory pathologies. The peptide, composed by leucines interrupted by lysine every four residues, was synthesized to simulate C-terminal amphipathic helical segments of SP-B. Once incorporated into lipid-protein complexes, KL4 promotes formation of interfacial films that produce and maintain surface tensions below 5 mN/m during compression-expansion cycling. Although KL4 was designed as an amphipathic helix at the membrane surface, the data on orientation and interactions of the peptide in membranes are contradictory. In the present work the surface activity and interaction with membranes of KL4 was compared with the behavior of two other peptides, KL2A2 and KL4PQ, to gain information about the properties defining the potential of KL4 as surfactant additive.The surface activity of the different peptides in lipid suspensions mimicking surfactant composition (DPPC/POPC/POPG 50:25:15 w/w/w), was analyzed in a captive bubble surfactometer (CBS). Only lipid suspensions containing KL4 adsorbed to the air-liquid interface with kinetics comparable to those containing SP-B, with KL4PQ and specially KL2A2 showing less activity. The interaction of the three peptides with membranes was analyzed by DSC, Laurdan fluorescence and ATR-FTIR. Perturbations induced by peptides on the structure and permeability of giant unilamellar vesicles (GUVs) were also analyzed and compared with the behavior of natural SP-B.The results indicate that the extent of penetration of the peptides in bilayers and their ability to perturb membrane structure correlate with their different interfacial activity.

Structure-Function Relationships of Antimicrobial Piscidins 1 and 3 bound to Cholesterol-Containing Lipid Membranes

Isolated in the mast cells of hybrid-striped sea bass, piscidin 1 (p1) and piscidin 3 (p3) are antimicrobial, cationic, and amphipathic peptides that have demonstrated broad-spectrum activity against bacteria, fungi, viruses, and cancer cells. Both p1 and p3 adopt an alpha-helical structure when bound to phospholipid membranes. p1 is the more active of the two isoforms, exhibiting higher lytic activity on different bacterial strains, as well as erythrocytes. This research uses various biophysical methods to investigate the differences in the backbone structure and bilayer location of piscidin bound to different lipid bilayers of biological relevance.To mimic the composition of human erythrocytes, a mixture of 4:1 palmitoyl-oleoyl-phosphatidylcholine (POPC):cholesterol (CHL) at pH 7.4 was used. Oriented bilayer samples prepared with 1:40 peptide:lipid were analyzed using solid-state NMR yielding 15N-1H dipolar couplings as well as 15N amide chemical shifts. These data were used to compute a high-resolution atomic-level structure of p1 and p3 bound to zwitterionic bilayers. The peptide's backbone structure is largely conserved across various bacterial and mammalian membrane mimics, suggesting that the difference in a peptide's activity on various membranes is more reliant on the peptide's side-chain and its position within the membrane.The structural results were supplemented with dye leakage assays in order to probe the peptide's activity on lipid vesicles, mimicking bacterial and mammalian cells. These results lend insight into how specific changes in peptide's structure may result in varying activity on different membrane systems. Molecular dynamics (MD) simulations were used to predict the peptide's bilayer location and depth of insertion. Membrane thinning, an event known to precede pore formation, was shown using MD. Overall, these experiments help obtain principles to design novel antibiotic pharmaceuticals with low hemolytic effects and high lytic activity on bacteria.

Structural Determinants for the Membrane Insertion of the Transmembrane Peptide of Hemagglutinin from Influenza Virus

Membrane fusion is a process involved in a high range of biological functions, going from viral infections to neurotransmitter release. Fusogenic proteins increase the slow rate of fusion by coupling energetically downhill conformational changes of the protein to the kinetically unfavorable fusion of the membrane lipid bilayers. Hemagglutinin is an example of a fusogenic protein, which promotes the fusion of the membrane of the influenza virus with the membrane of the target cell. The N-terminus of the HA2 subunit of this protein contains a fusion domain described to act as a destabilizer of the target membrane bilayers, leading eventually to a full fusion of the two membranes. On the other hand, the C-terminus of the same subunit contains a helical transmembrane domain which was initially described to act as the anchor of the protein to the membrane of the virus. However, in recent years the study of this peptide segment has been gaining more attention since it has also been described to be involved in the membrane fusion process. Yet, the structural characterization of the interaction of such a protein domain with membrane lipids is still very limited. Therefore, in this work, we present a study of this transmembrane peptide domain in the presence of DMPC membrane bilayers, and we evaluate the effect of several mutations, and the effect of peptide oligomerization in this interaction process. Our results allowed us to identify and confirm amino acid residue motifs that seem to regulate the interaction between the segment peptide and membrane bilayers. Besides these sequence requirements, we have also identified length and tilt requirements that ultimately contribute to the hydrophobic matching between the peptide and the membrane. Additionally, we looked at the association of several transmembrane peptide segments and evaluated their direct interaction and stability inside a membrane bilayer. From our results we could conclude that three independent TM peptide segments arrange themselves in a parallel arrangement, very similarly to what is observed for the C-terminal regions of the hemagglutinin crystallographic structure of the protein, to where the segments are attached.

Formation and domain partitioning of H-ras peptide nanoclusters: effects of peptide concentration and lipid composition.

Experiments have shown that homologous Ras proteins containing different lipid modification, which is required for membrane binding, form nonoverlapping nanoclusters on the plasma membrane. However, the physical basis for clustering and lateral organization remains poorly understood. We have begun to tackle this issue using coarse-grained molecular dynamics simulations of the H-ras lipid anchor (tH), a triply lipid-modified heptapeptide embedded in a domain-forming mixed lipid bilayer [Janosi L. et al. Proc. Natl. Acad. Sci. U.S.A. 2012, 109, 8097]. Here we use the same simulation approach to investigate the effect of peptide concentration and bilayer composition on the clustering and lateral distribution of tH. We found no major difference in the clustering behavior of tH above a certain concentration. However, the simulations predict the existence of a critical concentration below which tH does not form nanoclusters. Moreover, our data demonstrate that cholesterol enhances the stability of tH nanoclusters but is not required for their formation. Finally, analyses of peptide distributions and partition free energies allowed us to quantitatively describe how clustering facilitates the accumulation of tH at the interface between ordered and disordered domains of the simulated bilayer systems. These thermodynamic insights represent some of the key elements for a comprehensive understanding of the molecular basis for the formation and stability of Ras signaling platforms.

Solution NMR studies of peptide-lipid interactions in model membranes

Many important processes in life take place in or around the cell membranes. Lipids have different properties regarding their membrane-forming capacities, their mobility, shape, size and surface charge, and all of these factors influence the way that proteins and peptides interact with the membrane. In order for us to correctly understand these interactions, we need to be able to study all aspects of the interplay between lipids and peptides and proteins. Solution-state NMR offers a somewhat unique possibility to investigate structure, dynamics and location of proteins and peptides in bilayers. This review focuses on solution NMR as a tool for investigating peptide-lipid interaction, and special attention is given to the various membrane mimetics that are used to model the membrane. Examples from the field of cell-penetrating peptides and their lipid interactions will be given. The importance of studying lipid and peptide dynamics, which reflect on the effect that peptides have on bilayers, is highlighted, and in this respect, also the need for realistic membrane models.

How Lipid and Water Physical Chemistry May Control Fusion: Behavior of Membrane Interfaces and Fusion Peptides

Membrane fusion is an essential process in infection by enveloped viruses, synaptic exocytosis, and vesicle trafficking. As interacting membranes are brought together prior to fusion, the resulting membrane-water-membrane interface has surprising and non-bulk-like properties. Here we ask two questions regarding this membrane-membrane interface: how does the physical chemistry differ from bulk, and how do fusion peptides affect this interface. We have used atomistic molecular dynamics simulation to study these interfacial behavior at high resolution. At vesicle-vesicle interfaces prior to fusion, we observe in our simulations a distinct ordering of water between the two membranes and a concomitant slowing of dynamics. We used committor analysis to examine the role of fine water structure in fusion; holding the vesicles fixed while resampling water conformations from a solvent box substantially speeds fusion dynamics, with a small but significant effect on fusion energetics. These slowed dynamics may provide another reason for the reported propensity of fusion stalks to form around the periphery of a contact interface.We have also examined the effect of influenza fusion peptides in bilayers and at membrane interfaces. We and others have previously hypothesized that lipid tail protrusion is a precursor to fusion stalk formation. In our simulations, influenza fusion peptides promote tail protrusion most strongly in a kinked helix conformation with the ends inserted into the hydrophobic layer. Straight helix and helical hairpin conformations occupying an interfacial position promote tail protrusion less strongly.

Membrane Interactions of an Acylated and Non-Acylated Lactoferricin Peptide by Solid-State NMR and Fluorescence Spectroscopy and Molecular Dynamics Simulations

LfB6 (RRWQWR-NH2) is a tryptophan- and arginine-rich cationic antimicrobial peptide, derived from bovine lactoferrin, with broad spectrum activity that can be enhanced by N-terminal acylation (CH3(CH2)4CO-RRWQWR-NH2; C6-LfB6). The arginines promote selective interaction with negative charges present in bacterial cell membranes, while the tryptophans prefer the membrane-water interface. We have combined solid-state 2H and 31P NMR and fluorescence experiments with all-atom and coarse-grained molecular dynamics simulations to investigate the interactions of LfB6 and C6-LfB6 peptides in bilayers composed of two distinct compositions: 3:1 POPE:POPG (anionic) and POPC (zwitterionic). Previously we reported that the arginines of C6-LfB6 are first to associate with POPE:POPG; whereas in POPC the C6 tail associates first. Solid-state 2H NMR experimental results confirmed that the lipid order parameters are not significantly changed when C6-LfB6 is bound to negatively-charged membranes, while a slight decrease in order is observed for zwitterionic membranes (Romo, et al. 2011. Biochim. Biophys. Acta. 1808(8):2019-30). We now compare these results with those for non-acylated LfB6 peptide. Solid-state 2H NMR spectra of mechanically aligned samples reveal that the order of POPC and of POPG is reduced in the presence of 1 mol% LfB6. By contrast, POPE shows an increase in order and some line broadening. 31P NMR spectra indicate little effect on the lipid head group for either peptide. The Trp fluorescence emission maxima are blue-shifted for LfB6 (340 nm) and C6-LfB6 (336 nm) in POPE:POPG, compared to POPC (353 nm LfB6 and 348 nm C6-LfB6), suggesting that the Trp residues are less water-exposed in the anionic lipid membranes and when the C6 acyl chain is present.

Characterization of Antimicrobial Peptides Relating to Shortened RWALP Model Peptides

In the face of increasing bacterial drug resistance, small membrane-active peptides offer potentially attractive avenues to alternative antimicrobial agents. The goals of this project are to characterize the lipid interactions and analyze the antimicrobial efficacy of model peptides that are longer than the lactoferrin-derived, surface-acting LfB6 (RRWQWR-NH2), yet shorter than the transmembrane RWALP23 (acetyl-GRALW(LA)6LWLARA-NH2). New-generation RWALP peptides, of general sequence (ac-GRnWm(LA)jLWmRnA-NH2) were designed with varying total lengths (13-15) and numbers of Arg and Trp residues. 2H-alanines were incorporated at several positions to serve as probes for recording solid-state NMR spectra from mechanically aligned samples of the peptides in bilayers of DLPC, DMPC or DOPC. Labeled RWALP13 (j=3, n=1, m=1) exhibits partial water solubility, no antimicrobial activity, and 2H-NMR spectra characteristic of isotropic motion, even in the presence of lipid bilayers. Circular dichroism spectra of RWALP13 suggest partial α-helical character in water that is enhanced when lipids are present. The 2H-NMR spectra indicate that the longer 14- or 15-residue peptides are aligned to varying degrees in the different lipid bilayer membranes. Antimicrobial assays reveal that peptides with four arginines have higher activity than those with only two arginines. Interestingly, within the 4-Arg category, RRWALP15 (j=3, n=2, m=1) shows higher activity (MIC of 6.25 μ1/4g/ml) against E. coli than does RRWWALP15 (j=2, n=2, m=2; MIC of 25 μ1/4g/ml). The 2H-NMR spectra of RRWALP15 suggest significant alignment of the peptide with respect to lipid bilayer membranes. The combined antimicrobial and spectral features make RRWALP15 an especially good candidate for further analysis.