Structure and mechanism of human antimicrobial … · Structure and mechanism of human antimicrobial peptide dermcidin and its antimicrobial potential Kornelius Zeth1,2 1Unidad de

Structure and mechanism of human antimicrobial peptide dermcidin and its antimicrobial potential

Kornelius Zeth1,2 1Unidad de Biofisica (CSIC-UPV/EHU) and Fundación Biofísica Bizkaia, Barrio Sarriena s/n, 48940, Leioa, Vizcaya,

Spain 2IKERBASQUE, Basque Foundation for Science, Bilbao, Spain.

Human beings control viral, bacterial and fungal infections by producing peptide-derived broad-spectrum antibiotics. This battery of host defense molecules can compromise the integrity of microbial cell membranes and thereby evade various pathways by which bacteria develop rapid antibiotic resistance through mutational adaption. Although more than 2000 antimicrobial peptides (AMPs) from various species have been described and some of which are studied in great detail, the structural and mechanistic basis of their activity remains largely unknown. To understand the mechanisms by which AMPs compromise cell membranes we performed structure biology, electrophysiology and simulations on human dermcidin (DCD) in artificial membranes. Together these techniques reveal the antibiotic mechanism of this fascinating channel structure. The structure of DCD is the first AMP tracked in an oligomeric channel configuration in solution. DCD forms a novel architecture of high-conductance transmembrane channels, composed of zinc-stabilized anion-selective hexameric pores. Molecular dynamics simulations elucidate an unusual membrane permeation pathway for ions and show adjustment of the pore to various membranes by variable tilting angles. Our study comprehensively unravels the mechanism for membrane-disruptive action of this particular mammalian host defense peptide.

Keywords Antimicrobial peptides; human epithel; Dermcidin; channel structure; barrel stave model, electrophysiology

1. Introduction

The danger of many bacteria becoming more resistant to traditional antibiotics has recently been placed on the national risk register of many European countries and the US, ensuring that it will be given full attention by the World Health Organization (WHO) [1]. These warnings are in consequence of the steep global rise in infections by multi-resistant bacteria also known as ‘superbugs’ also known as ‘ESKAPE’ group (Enterococcus spp., Staphylococcus aureus, Klebsiella spp., Acinetobacter baumannii, Pseudomonas aeruginosa and Enterobacter spp.). Indeed, among the critical cases of some superbugs are e.g. the carbapenem-resistant enterobactericeae (CRE), methicillin-resistant Staphylococcus aureus (MRSA) strains, Neisseria gonorrhoeae and entirely resistant Acinetobacter baumanii bugs [2-10]. A recent USA today report summarizing hundreds of scientific papers claims that the CRE strains of Klebsiella pneumoniae recently battered several US hospitals among which were the National Institutes of Health (NIH) with seven dead people [2,6]. Death rates among patients carrying CRE or A. baumanii are about 40%, far worse than e.g. statistics of MRSA cases reported in the past [2,5,6,9]. The ´superbugs´ develop rapid resistance most significantly in association with hospitals running at low hygienic standards and patients treated inappropriately with high doses of antibiotics. They most often target humans already compromised in their immune system while healthy individuals are well protected by an intact innate and adaptive immune system. This strong dissemination in severe and untreatable infections gives rise to speculations that even routine hospital operations could become much more risky within several years [1]. Currently there is little hope that an effective drug to kill these superbugs will be developed within the next years due to the little incentive for pharmaceutical companies competing with the rapid adaption of these bacteria [1]. In the light of the alarming development we find one evolutionary ancient success story behind the survival of mammals given the high power of different microbes: the innate immune system [11,12]. This sophisticated response system producing numerous physical and chemical barriers allowed humans to have successfully survived many of those threads. Multicellular organisms fight bacterial and fungal infections by producing mucus, low pH conditions (in the stomach) and specific blood proteins, fighting for iron as an essential nutrient for all bacteria [13]. Against the background of growing concern about untreatable infections, there are hopes that especially a new generation of antibiotics based on natural peptides can be further developed before certain infections become untreatable. All higher organisms produce peptide-derived broad-spectrum antibiotics as their second line of defense [14-18]. These constantly expressed peptides allow not only for the controlled and ´friendly´ co-existence of bacteria and host species, but also keep the number of about 1015 bacteria accompanied with e.g. human beings at a tolerable upper boundary. They also suppress infections at an early stage of their development through an inducible concentration enhancement gradient around new wounds [19]. These host defense peptides most often compromise the integrity of microbial cell membranes and thus evade mechanisms of genetic adaption by which bacteria develop rapid antibiotic resistance [14-18]. Traditional antibiotics are mostly targeting a small set of essential enzymatic pathways such as (I) iron uptake, (II) peptidoglycane synthesis, (III) protein biosynthesis or (IV) DNA replication and are studied in great detail [20-24]. While, a large body of information on the structure and function of AMPs has been accumulated, their mechanistic

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description rely on the continuous refinement of essentially three different models in the literature using indirect methods of analysis: the barrel-stave mechanism, the carpet and toroidal model (see Figure 1). In comparison to small-molecule antibiotics, host defense or antimicrobial peptides are often considered to have a distinctly superior properties, as they target the ‘microbial Achilles heel’, i.e. the unique but essential features of all microbial cellular membranes (albeit at somewhat lower efficacy) [16]. The drawback of this unique targeting mechanism causes a lack of knowledge on AMP killing mechanisms because the bacterial cell membrane and peptides structures embedded therein are more difficult to study and the problems are related to the difficulties of studies on membrane proteins (in comparison to soluble proteins). Studying the active and killing conformations of peptides in membranes, however, are important next steps to be discovered in help of the development of the desired rational approach of these agents further into next-generation antibiotics.

2. Mechanisms of antimicrobial peptides targeting membranes – barrel stave, toroidal and carpet model

General models for peptide membrane interactions are developed on the basis of mostly indirect experimental results and theoretical modeling using artificial membranes resembling the cytoplasmic membrane of Gram-positive and negative bacteria. The models developed in the literature assume the presence of a α-helical amphipatic peptide interacting with membranes while the mechanism by which β-sheet AMPs interact with cells is not yet integrated [17,25]. Peptide concentrations in solution also govern the interaction between artificial and biological membranes. At low concentrations peptides interact (positively charged/hydrophobic) with the negatively charged lipid head groups based on their amphipatic structure and may increase the local concentration of peptides through clustering. Increasing the concentration might shift the equilibrium between monomeric, dimeric and oligomeric states already in solution. At higher concentrations and potentially driven by an external electric field or an electrical gradient over a biological membrane these peptides may execute membrane permeabilization following one of the three models. Interestingly, also some pore-forming bacteriocins such as Colicin A, IA or S4 have been described to target the inner bacterial membrane or lipid bilayers only in response to a membrane potential [26-28]. Due to the functional analogy of pore formation between bacteriocins and antimicrobial peptides it appears plausible, that pore formation in membranes occurs after pre-assembly and voltage dependent insertion (e.g. for DCD). In the barrel-stave model amphipatic peptides are vertically oriented to the membrane facing the lipidic environment charged side chains [25]. This transmembrane pore based on the barrel stave model has been studied on the basis of experimental Alamethicin oligomerization data in membranes. Several oligomerization models of 3-11 molecules of Alamethicin have been reported yielding an inner diameter of ~ 2-4 nm [29,30]. More recently a tetrameric arrangement of parallel peptides in membranes has been proposed on the basis of electron paramagnetic resonance data and molecular dynamic simulations [31]. In the carpet model amphipatic peptides are initially oriented in parallel relative to the membrane surface of the lipid layer [25]. Increasing the concentration of the peptide eventually causes small lipidic droplets of micellar shape to be formed, surrounded by amphipatic peptides. Although this arrangement appears plausible, it requires a large number of peptides to cover patches of the bacterial membrane. Finally, the membrane is significantly disturbed after the release of peptide coated micellar lipid complex. One of the prominent representatives of this model is melittin, a 26 residue long peptide from venom and honeybees. This peptide has recently been demonstrated to form pores in the outer and inner membrane of Gram-negative bacteria flowed by the efflux of protein from the cytoplasma [32]. The third model, the toroidal pore model is similar to the barrel stave model in that both arrangements form pores in artificial and biological membranes, respectively. By contrast to the barrel stave model, here lipid head groups contribute to the pore formation and stabilization. Two well studied representatives of the mode of pore formation are Magainin 1 and 2 [29,33,34].


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Fig. 1 The models currently used to describe the interactions of antimicrobial peptides with artificial and natural membranes. The carpet model shown in (A) assumes an initial interaction of amphipatic peptides with the lipid bilayer surface driven by an electrostatic attraction between positively charged peptides and negatively charged lipid bilayers. The driving force for the peptides to align vertically instead of horizontally may be introduced by application of a transmembrane potential e.g. an inner membrane potential. (B) The toroidal is similar to the barrel stave model in the assumption of a spatially defined pore. However, while the barrel stave pore is assumed to be exclusively built by peptides, the toroidal pore may be stabilized in addition to peptide-peptide contacts by lipid head group interactions. (C) The carpet model deviates from the previous models assuming that part of the lipids undergo a phase transition from a bilayer to a rather micellar phase stabilized by peptides in the presence of large peptide concentrations. After the formation of these peptide-stabilized micelles, individual complexes may diffuse apart from the membrane to leave holes in their back.

3. Antimicrobial peptides protect from bacterial overgrowth of the human body

Host defense peptides actively control a wide range of microbes across tissues of the animal and plant kingdoms, which signify their importance during the evolution of multicellular organisms [16,17]. The size of these mostly amphipatic and positively charged peptides varies between 10-50 residues. The human body expresses several anti-microbial peptides (hAMPs) as part of the ancient innate immune system. These mostly cationic peptides are classified into alpha-, beta-defensins and histatins which form the largest subgroups of hAMPs, while cathelicidin (LL-37) and dermcidin (DCD) are additional single representatives of this structurally diverse group [35,36]. All peptides are initially produced as inactive precursor proteins comprising pre-sequences, subsequently delivered as inactive form to their destination where they become activated through proteolysis [36]. Local proteases e.g. on the skin typically produce a mixture of peptides derived from the full-length peptide in different lengths which were shown to often also harbor a varying activity against a broad variety of microbes tested. The epidermal surface e.g. forms a two m2 large surface which is occupied by bacterial species from the Staphylococcus and Streptococcus groups. Peptide mixtures of DCD, LL-37 and others prevent continuous infections due to lesions on the skin surface at low concentrations in the 5-30 μg/ml range and coincide with the MIC values determined for LL-37 and DCD [35,36]. The general scheme of peptide processing is shown using DCD as the example (see Fig. 2).


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Fig. 2 The precursor protein of the dermcidin peptide is produced in sweat glands comprising a signal peptide, a pro-peptide and the AMP domain. Pro-peptide and AMP domain are delivered to the skin surface and cleaved by proteases releasing the pro-peptide from the mature AMP domain. The sequence of the AMP domain is shown in a box with positively charged residues marked in blue and negatively charged residues in red yielding the overall charge of the domain to be negative. This domain and fragments of it are actively killing bacteria.

4. The crystal structure of the human Dermcidin channel

Chemically synthesized DCD was crystallized in the presence of zinc atoms but in the absence of detergents or lipids and the structure was solved by single anomalous dispersion [37]. The structure of the DCD assembly at 2.5 Å resolution exhibits a novel peptide channel architecture comprising a hexameric bundle formed by elongated α-helices, which adopts overall dimensions of ~8 × 3.5 nm2 (Fig. 3A). The helices are essentially ideal 413-helices without any supercoil contribution and align well with many single amphipathic helical peptides of the PDB database. Channel formation involves trimerization of anti-parallel peptide dimers resulting in a firmly enclosed channel structure (see Fig. 3A). Each peptide monomer touches two distinct interfaces to neighboring subunits (see Fig. 3A). The more extended interface, displaying a contact surface of 930 Å2, is mainly stabilized by hydrophobic interactions, while the second interface covers only 520 Å2 (IF1) and is primarily stabilized by Zn2+ ions that intercalate between the two helix ends (Fig. 3A). The zinc binding in DCD is coordinated by N- and C- terminal residues of dimerizing peptides (residues involved: Glu5, Glu9, His38’, Asp42’; Fig. 3A, B). They are exposed to the inner wall of the channel and substantially change the overall charge of the assembly (from -12e counting charged residues to neutral). In addition, they modify the local charge distribution in particular at the entrance of the channel. A second zinc binding site S2 was identified exposed to the channel surface where zinc ions are ligated by two Asp residues 24 and 28. Assuming the peptide complex is integrated in the membrane this Zn2+ binding site would face the membrane interior and therefore is likely non-physiological. Moreover, the S2 sites are unusual binding sites providing only two residues for zinc chelating in contrast to the S1 site architecture resembling a typical binding site for divalent ions with the ion kept in a hexagonal complex arrangement.


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Fig. 3 Structure of dermcidin shown in surface and cartoon representation as side and top view and as zoom-in into the zinc binding site. (A) The surface representation of DCD is shown with the individual peptides color coded in orange (α) and blue (α∗). The two interfaces of one peptide are marked in with a dashed yellow line and termed IF1 and IF2. In the middle the hexameric complex in cartoon representation is shown. The length of the peptide complex is given in nm and the zinc binding sites S1 and S2 are marked with rectangles. On the right hand side the complex is shown along the axis and the outer diameter is given in nm. The threefold axis of the complex is marked with a triangle and termed C3. (B) Zoom-in into the zinc binding site 1 (S1). Residues involved in co-factor binding are marked accordingly and the distance between the two binding sites is given in nm. (C) The second and presumably artificial zinc binding site (S2) is shown with the residues involved in binding highlighted in stick representation. All structure pictures were prepared using PYMOL [38]. DCD shows a unique distribution of charges, reflecting a peptide channel that occurs in both, soluble and membrane bound forms. The entire hexameric channel comprises 96 ionizable residues, which are all oriented toward the channel interior (Fig. 4A). This enormous charge density, which is not completely shielded to the exterior, is likely to contribute to the relatively high aqueous solubility of DCD. The inner space of the peptide channel has an apparent separation into five radially symmetric charge girdles I/II/III/II/I (Fig. 4A). Residues presumably facing the acyl chain region of the membrane are exclusively hydrophobic (Ala, Val, Leu) and the channel has the surface properties of a pure α-helical membrane protein (Fig. 4B). However, no aromatic residues are found that are commonly considered to be important for the lateral adjustment of proteins in membranes [39,40]. The channel diameter is not homogeneous and varies along the y-axis with two rather narrow entry sites, followed by a widened interior with ‘window’-like eyelets in the IF1 interface (Fig. 4C). The six lateral openings have a diameter of ~1 nm and are surrounded by small amino acids as well as positively charged residues which may have an influence on the selection of ion entry as the channel was determined to be anion selective. The distance between two adjacent eyelets is 2.5 (in same plane) and 3 nm (in the opposite plane), respectively, roughly corresponding to the diameter of the membrane hydrophobic core [37].


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Fig. 4 Analysis of dermcidin surface properties including charges and amphipatic patterns. (A) Surface representation of the DCD hexamer with two helices (shown in orange and blue) are omitted from the surface calculation. Negatively charged surface patches are color coded in red, positively charged surface patches are color coded in blue. Water molecules from the crystal structure are marked as blue balls. (B) Surface representation of the entire hexameric channel with hydrophobic residues facing the surrounding is marked in magenta and residues are marked with type and number. (C) Representation of DCD helices in cartoon design together with a surface representation of the ‘empty space’ inside the channel. There are two different entrance/exit opportunities from the termini (two) of the structure and from the side (six). The crystal structure of DCD provides the first glimpse into the true oligomeric assembly of peptides building a peptide channel in an active conformation. Additional experimental data on DCD have been collected by other groups whereby a structure of DCD has been determined by high resolution NMR studies carried out in 50% TFE and 100 mM SDS [41]. These experimental conditions are chemically and physiologically far away from the pure solvent exposed or membrane integrated state determined by X-ray crystallography and hence may introduce serious disorder in the molecule arrangement. In the monomeric NMR structure (PDB-entry 2KSG) four small helices connected by β-turns are modeled to reflect the experimental parameters. This model determined under non-physiological conditions does not resemble secondary structure predictions using PSIPRED or JPRED servers and also lacks any structure analogy to the hexameric crystal structure determined in a detergent-free state [42,43]. More recent experiments aiming to understand DCD assembly and oligomerization were published by the Schittek group. In a paper by Paulmann et al. the authors describe two ‘novel’ DCD properties: (I) the influence of zinc ions on the oligomerization state along with (II) a putative model in planar lipid bilayers. Peptide interactions with zinc as a hypothetical cofactor were miraculously ‘guessed’ without supporting experimental evidence. Furthermore, neither the effect of zinc on the activation of DCD, although postulated, was proven by their experiments using NMR and electrophysiology. Consequently the model developed on the basis of vague experimental data is wrong. In this model a putative kink of the elongated helices and the parallel orientation stand in strong contrast to our own observations and are unlikely the results of experiments. Finally, the strong effect of zinc on the channel function (although tested in their paper) was overseen by the Schittek group [44].

5. Functional properties of the DCD channel in the presence and absence of the zinc co-factor

When we had determined the channel structure a number of questions turned up regarding the physiological relevance of the channel conformation and the putative mechanism of function. So we were wondering if the channel conformation seen in crystallography based on high peptide (mM range) and zinc concentrations (100 mM zinc) are relevant also at low concentrations using planar membrane technology. Under these conditions two reservoirs are filled with a 1 M NaCl solution and separated by a single lipid bilayer. To obtain functional evidence for the activity of DCD channels in the presence and absence of zinc, we collected conductivity data in these planar lipid bilayers. The oligomeric peptide in the absence of zinc introduced conductivity over the membrane which showed a noisy behaviour of the channel resulting in a conductance of ~35 pS. In the presence of zinc the activity of the peptide was much higher and reliable statistics could be obtained using > 1000 insertions or gating events, respectively. The conductivity determined under these conditions was significantly higher with ~80 pS at a significant open probability (see Fig. 5B and 5C). These experimental data support the findings from X-ray crystallography of a channel which, in the presence of zinc ions, resembles stable, specific and defined individual oligomer with a high conductance. Although speculative, the conductance data plausibly indicate two different conformational states of the channel in the membrane which were previously being modelled by MD techniques using the peptide structure in the presence and absence of zinc. From these modelling data it became obvious that the interface 2 which is stabilized by H-bonds and hydrophobic interactions


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remains stable, the second interface of the monomer lost stability in the absence of zinc and the peptide dimer pairs drifted away from each other [37].

Fig. 5 Electrophysiology performed on the DCD channel in the presence and absence of zinc ions. (A) Current trace of DCD peptide channels at a transmembrane potential of +100 mV. Protein activity was only rarely observed under these conditions. (B) Current traces of DCD recorded at a holding potential of +100 mV in the presence of zinc. The current traces clearly show a defined conductivity state of approx. 80 pS. (C) Increasing the concentration of DCD lead to the insertion of more than one channel (in this case two) with the doubled conductivity level as determined for the monomer. (D) Molecular dynamic studies of DCD in solution and in the absence (top) and presence (bottom) of zinc. The study indicates the putative role in stabilizing the hexamer at the interface I (IF1) while this interface becomes distorted in the absence of zinc. Notably, the hexamer remains stable as a complex but the termini are bent apart from each other.

6. Molecular dynamic studies provide insights into a fascinating and novel ion translocation mechanism

In order to obtain a mechanistic picture of DCD embedded in artificial membranes. We performed atomistic molecular dynamics (MD) simulations of the channel integrated into negatively-charged, bacterial-like phosphatidyl ethanolamine/ phosphatidyl glycerol bilayers [37]. These simulations were initiated to follow the ion permeation properties of the channel. The peptide assembly including Zn2+ ions exhibited a high level of structural stability in membranes. After initially positioning the complex normal to the bilayer plane, the assembly adopted tilted configurations with angles of 20-30 degrees relative to the membrane. The inclination is likely to compensate for the hydrophobic mismatch between the bilayer and the hydrophobic exposed surface, and its magnitude depended on the length of the surrounding lipids.

Fig. 6 Molecular dynamic studies of DCD suggest tilting of the channel in artificial membranes and an unexpected path for anion translocation. (A) Molecular dynamic trajectories of DCD (md1, md2 and md3) all lead into the same tilting angle of ~ 20-30 degrees relative to the membrane normal. (B) The channel in surface representation and the lipids in stick representation are shown from top. Water molecules inside the channel are shown in red. (C) The channel in a side view with the lipid matrix indicated as surface representation in grey. Zinc ions in grey and chloride ions are marked and their way of diffusion from the lateral space is indicated by arrows.


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During the simulations, the inner sphere of the oligomeric complex is filled with water molecules and developed a permanent water channel across the membrane (Fig. 6B). To investigate the conductivity of DCD for ions, we used computational electrophysiology to model biologically realistic electrochemical gradients across membranes. At 1 M NaCl concentration, which is the concentration being used for the experimental determination of conductivities, we obtained a channel conductance of ~110 pS, in excellent agreement with our experimental data. In simulations at a salt concentration of 150 mM, a total single channel conductance of ~50 pS was obtained (Fig. 6A). All simulations of DCD showed pronounced anion selectivity. The remarkable agreement between computational and experimental data strongly suggests that the hexameric crystal structure is identical to the functional state in microbial membranes. Any higher or lower oligomerization state of DCD would very likely lead to drastically different conductance values. This interpretation is corroborated by the strong dependence of channel current on zinc, which we observed in both the electrophysiology experiments and MD simulations in membranes, and which corresponds directly with the abundance and function of the Zn2+ binding sites, linking the channel subunits in the crystal structure. Notably, the DCD channel exhibited a unique and unexpected ion permeation pathway in the simulations, which offers an explanation for this unexpectedly high conductance, in spite of its limited channel cross section. Through channel tilt, ions are capable of entering sideways into the pore across the eyelets that occur at the trimeric interfaces. This not only shortens the pathway across the channel considerably, but more importantly, exploits the increased ion concentration observed at the lipid head groups by enabling these ions to enter the channel directly, and to rapidly traverse the inner pore (Fig. 6C). Also inside the channel, DCD shows an unusual anion traversal mechanism. Most anion transfer steps across the inner section of the pore consist of single ion 'hopping' transitions. Near the channel termini, however, anions accumulate to form clusters of 3-4 ions, most clearly seen at the channel exit. Productive ion translocations exiting the channel usually involve multi-ion “knock-on” effects, through which individual anions are expelled from this cluster to the bulk solution.

7. Dermcidin shows a deviation of the barrel stave mechanism

The experimental data collected on dermcidin propose a modified mechanism based on the barrel stave model (see Fig. 7). In contrast to this original model which assumes, that isolated peptides attach to the membrane resulting from electrostatic attraction, we observed, that the channel is a stable hexamer in solution and membranes. This finding is important for our understanding of the barrel stave mechanism and suggests a preassembly of the channel in solution. Given the large membrane surface of a bacterium and the limited number of peptides attached to the membrane, this early oligomerization step increases the probability of peptide association and channel formation significantly. Thereby the likelihood of channel formation in membranes followed by killing bacteria is strongly increased. Our unpublished data of DCD imply a strong tendency of this channel to remain in solution or to attach to artificial lipid surfaces without further insertion, unless a membrane potential of approx. 100 mV is applied [37].

Fig. 7 Model of DCD insertion into artificial/biological membranes. Model of DCD interactions with membranes. Peptide monomers of DCD are in equilibrium with the hexameric channel form, which is stabilized by the presence of zinc. The channel and DCD monomers can interact with membrane surfaces through lateral association as shown by our NMR studies. The channel can be translocated into the membrane upon application of a TMV, as shown in planar lipid membrane conductance measurements, and forms a representative of the barrel-stave model. This process may resemble the TMV of bacterial or fungal membranes. Once the channel is integrated into a membrane, channel conductance can be recorded by planar lipid membrane experiments and estimated by molecular dynamics studies. By contrast to the previously proposed barrel stave model, this model is based on the finding that DCD is an oligomeric complex in solution and membrane diffusion step as assumed for many AMPs are negligible.


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8. Dermcidin – a potential antibiotic?

Peptides are generally well represented in pharmacological studies and are the basis of a billion euro market including e.g. peptides added in food and as supplement in beauty crèmes. However, these peptides are either small and/or chemically stabilized through the introduction of non-natural amino acids or cystein disulfide bridge crosslinking. While the concept of antimicrobial peptides as powerful pharmaceuticals in the treatment of bacterial and fungal infection is well established, their current use is hampered by a number of factors. One and possibly the most important factor is the problem to synthesize/produce these peptides in large amounts given the high cost of chemical synthesis proportional to peptide length. Further problems are their stability under in vitro conditions where high concentrations of proteases are present. These problems mostly occur when peptides are administered in the body either as oral medication or intravenous. Their application on the human tissue however is promising and peptides on the basis if the LL-37 sequence against e.g. chronical bacterial ear infection are in phase II and III trials [45]. Several patents demonstrate a growing interest of universities and companies such as B.R.A.I.N in the large scale production of the DCD peptide for future applications in pharmaceutical research [46,47]. However, there is still a long way to get more and successfully AM peptides into pharmaceutical pipelines and there are more problems to overcome than those mentioned on the way to become a pharmacologically interesting and rewarding target. Consequently, although DCD is a well-studied peptide and understood in molecular detail, it will take years to firmly investigate if it has the potential to become a lead substance in the treatment of bacterial infections on the skin.

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Structure and mechanism of human antimicrobial … · Structure and mechanism of human antimicrobial peptide dermcidin and its antimicrobial potential Kornelius Zeth1,2 1Unidad de