Current Topics in Medicinal Chemistry - Volume 16, Issue 1, 2016

Antimicrobial peptides (AMPs) establish the first line of host defense mechanism against invading microorganisms including bacteria, viruses, fungi and parasites. In recent years, emergence and spread of antibiotic resistance bacterial pathogens have dawn considerable interest in investigations of AMPs. The ability of AMPs to exert lethality against multiple drug-resistant (MDR) bacteria has incited promising avenues for antibiotic development. As a mode of action, most AMPs perturb the membrane organization of bacterial cells. The outer membrane lipopolysaccharide (LPS) of Gramnegative bacteria establishes a superior permeability barrier, in contrast to the peptidoglycan layer of Gram-positive bacteria. Due to LPS barrier, development of antibiotics for drug resistant Gram- negative bacteria are more complicated, with only fewer compounds in the pipeline. Recent studies have demonstrated that LPS actively regulate mode of action of AMPs on the lethality of Gram-negative bacteria. LPS, also known as endotoxin, is the primary agent for septic shock syndromes in intensive care unit killing over 120,000 people in the USA. Currently, anti-sepsis therapies are greatly lacking. Therefore, LPS has been considered as a target for the development of antimicrobial and antisepsis drugs. In recent and past few years, 3-D structures and interactions of a number of AMPs have been determined in complex with LPS micelles. These studies have generated molecular insights towards mode of action and synergistic activity of AMPs in the outer membrane. In this review, atomic resolution structures and interactions of potent AMPs with LPS are discussed providing novel insights of their mode of action.

Antimicrobial peptides (AMPs) have attracted considerable recent interest as potential therapeutics, motivated by increasing resistance development against conventional antibiotics. This brief overview summarizes some key aspects related to the interaction of AMPs with bacterial and cell membranes, as well as with membrane components, which is at the core of the mode-of-action of these compounds. Throughout, studies on peptide interactions with model lipid membranes and membrane components are correlated to biological results on antimicrobial and anti-inflammatory effects of AMPs, and translated into therapeutic considerations.

Antimicrobial peptides (AMPs) are showing increasing promise as potential candidate antibacterial drugs in the face of the rapidly emerging bacterial resistance to conventional antibiotics in recent years. The target of these peptides is the microbial membrane and there are numerous models to explain their mechanism of action ranging from pore formation to general membrane disruption. The interaction between the AMP and the target membrane is critical to the specificity and activity of these peptides. However, a precise understanding of the relationship between antimicrobial peptide structure and their cytolytic function in a range of organisms is still lacking. This is a result of the complex nature of the interactions of AMPs with the cell membrane, the mechanism of which can vary considerably between different classes of antimicrobia peptides. A wide range of biophysical techniques have been used to study the influence of a number of peptide and membrane properties on the cytolytic activity of these peptides in model membrane systems. Central to characterisation of this interaction is a quantitative analysis of the binding of peptide to the membrane and the coherent dynamic changes in membrane structure. Recently, dual polarization interferometry has been used to perform an in depth analysis of antimicrobial peptide induced membrane perturbation and with new mass-structure co-fitting kinetic analysis have allowed a real-time label free analysis of binding affinity and kinetics. We review these studies which describe multi-step mechanisms which are adopted by various AMPs in nature and may advance our approach to the development of a new generation of effective antimicrobial therapeutics.

Melittin is a 26 residue peptide and the major component of bee (Apis mellifera) venom. Although melittin has both anticancer and antimicrobial properties, utilization has been limited due to its high lytic activity against eukaryotic cells. The mechanism of this lytic activity remains unclear but several mechanisms have been proposed, including pore formation or a detergent like mechanism, which result in lysis of cell membranes. Several analogues of melittin have been synthesized to further understand the role of specific residues in its antimicrobial and lytic activity. Melittin analogues that have a proline residue substituted for an alanine, lysine or cysteine have been studied with both model membrane systems and living cells. These studies have revealed that the proline residue plays a critical role in antimicrobial activity and cytotoxicity. Analogues lacking the proline residue and dimers of these analogues displayed decreased cytotoxicity and minimum inhibition concentrations. Several mutant studies have shown that, when key substitutions are made, the resultant peptides have more activity in terms of pore formation than the native melittin. Designing analogues that retain antimicrobial and anticancer activity while minimizing haemolytic activity will be a promising way to utilize melittin as a potential therapeutic agent.

The essence of successful antimicrobial chemotherapy lies in selective toxicity of the agent towards the pathogen. An ideal antimicrobial agent should kill pathogens effectively with little or no effect on host cells. There is a dearth of antibiotic and antimicrobial therapies due the rapid development of microbial resistance to these agents, as evidenced by increasing incidences of hospital acquired infections. This challenge necessitates the discovery and development of novel and effective antimicrobial agents. One promising approach is Antimicrobial Peptides (AMPs), which are synthesized by a large number of organisms. The presence of AMPs throughout evolution hints at their importance. The first and foremost interaction between AMPs and target cell occurs at the membrane of the pathogen. The details about these interactions will pave way for the development of new synthetic analogues or modified analogues of existing AMPs. Mechanistic insights into adoption of different structures in presence of bacterial membranes (and with their specific targets) will enhance our understanding and knowledge about these agents and their detailed mechanism of action. AMPs interact with lipids and form lipid-AMP complexes that create AMP-lined ion channels, which in turn modulate the membrane potential. This may have an effect on various biological processes leading to arrest of cell growth or cell death. This review summarizes the ion channel formation property of AMPs as an effective approach in dealing with neutralization of pathogenic microbes.

The extensive search for alternative therapeutics against microbial pathogens has led to the discovery of cationic peptides as new anti-infectives with a novel mode of action. Particular interest has been devoted to small linear peptides that can be efficiently made by chemical synthesis at competitive costs. The most promising originate from a large family of short, naturally occurring peptides found in the skin of amphibia of Rana genus, i.e. the temporins. This review is mainly focused on the recent structure-function studies of the earliest known temporin isoforms (TA, TB and TL) and their potential clinical role as novel antimicrobial agents. The development of novel antibiotics is an urgent public health concern due to the increased resistance of microorganisms to conventional antibiotics, particularly in the hospital setting.

The structural requirements for the synergistic enhancement of antimicrobial activities of the cationic linear peptides PGLa and magainin 2 were investigated. In a first step the antimicrobial activities were evaluated for a number of modifications of the sequences and equimolar mixtures thereof. In particular fluorophore labelled peptides maintain a high degree of antimicrobial activity and considerable synergism when tested conjointly. Thereafter, circular dichroism spectroscopy indicated that these extended sequences adopt helical conformations in the presence of model membranes similar to the unmodified sequences. Energy transfer between the fluorophores suggested that the peptides reside in close proximity to each other when bound to the membrane surface at high concentrations. The fluorophore interactions quickly diminish at lower peptide-to-lipid ratios indicating that the peptide-peptide interactions are weak. Furthermore, 15N solid-state NMR measurements of the membrane topology of [15N-Ala14]-PGLa and of its fluorophorecarrying analogue reconstituted into supported 1, 2-didecanoyl-sn-glycero-3-phosphocholine bilayers were performed. These experiments revealed no correlation between the topological state of PGLa and the observed synergistic enhancement of antimicrobial activities due to the presence of magainins. These results suggest that lipid mediated interactions rather than the formation of tight peptide-peptide complexes in the membrane are responsible for synergistic activities of the mixtures.

Antimicrobial peptides (AMPs) are a large class of innate immunity effectors with a remarkable capacity to inactivate microorganisms. Their ability to kill bacteria by membranolytic effects has been well established. However, a lot of evidence points to alternative, non-lytic modes of action for a number of AMPs, which operate through interactions with specific molecular targets. It has been reported that non-membrane-permeabilizing AMPs can bind to and inhibit DNA, RNA or protein synthesis processes, inactivate essential intracellular enzymes, or affect membrane septum formation and cell wall synthesis. This minireview summarizes recent findings on these alternative, non-lytic modes of antimicrobial action with an emphasis to the experimental approaches used to clarify each step of their intracellular action, i.e. the cell penetration mechanism, intracellular localization and molecular mechanisms of antibacterial action. Despite the fact that such data exists for a large number of peptides, our analysis indicates that only for a small number of AMPs sufficient data have been collected to support a mode of action with an authentic and substantial contribution by intracellular targeting. In most cases, peptides with non-lytic features have not been thoroughly analyzed, or only a single aspect of their mode of action has been taken into consideration and therefore their mechanism of action can only be hypothesized. A more detailed knowledge of this class of AMPs would be important in the design of novel antibacterial agents against unexploited targets, endowed with the capacity to penetrate into pathogen cells and kill them from within.

The Antimicrobial peptides (e.g. defensins, hevein-like molecules and food-protecting peptides like nisin) are able to interact specifically with contact structures on pathogen surfaces. Besides protein receptors, important recognition points for such contacts are provided by pathogen glycan chains or surface lipids. Therefore, structural data concerning surface exposed glycans and lipids are of the highest clinical interest since these recognition functions play a key role when optimising anti-infection therapies. Approaches in nanomedicine and nanopharmacology in which various biophysical techniques such as NMR (Nuclear Magnetic Resonance), AFM (Atomic Force Microscopy), SPR (Surface Plasmon Resonance) and X-ray crystallography can be combined with biochemical and cell-biological methods will lead to improved antimicrobial peptides by this rational drug design approach. Such a strategy is extremely well suited to support clinical studies focussing on an effective fight against multiresistant pathogens. The data sets which are described here can be considered as universal for the design of various antimicrobial drugs against certain pathogens (bacteria, viruses and fungi) which cause severe diseases in humans and animals. Furthermore, these insights are also helpful for progressing developments in the field of food conservation and food preservation. A detailed analysis of the structure-function relationships between antimicrobial peptides and contact molecules on pathogen surfaces at the sub-molecular level will lead to a higher degree of specificity of antimicrobial peptides.

Antimicrobial peptides (AMPs) are integral components of the host innate immune system and functional throughout the plant and animal kingdoms. AMPs are short cationic molecules and lethal against a wide range of bacteria, viruses, fungi, yeast and protozoa due to their membranolytic effects on the negatively-charged microbial membranes. In addition, they exert multiple immunomodulatory roles like chemotaxis, modulation of cytokine and chemokine expression, leukocyte activation etc. Since AMPs suffer loss of microbicidal properties under serum and tissue environments, their capacity to modulate the immune system may predominates under the physiological conditions. Discovery of new antibiotics is lagging far behind the rapidly spreading drug resistance among the microorganisms. Both natural and synthetic AMPs have shown promise as ‘next generation antibiotics’ due to their unique mode of action, which minimises the chance of development of microbial resistance. In addition, they have therapeutic potential against non-infectious diseases like chronic inflammation and cancer. Many of the synthetic AMPs are currently undergoing clinical trials for the treatment of debilitating diseases, such as catheter-related infections, diabetic foot ulcers, chemotherapy-associated infections etc. Some of them have already entered the market as topical preparations. In this review, we synopsise the current literature of natural and synthetic AMPs in different infectious and inflammatory diseases of human microfloral habitats, especially the gastrointestinal, respiratory and genitourinary tracts and the skin. We also discuss the classification of AMPs, their mode action and antimicrobial spectrum, including the pathogen evasion mechanisms. In short, we tried to present the locus standi of AMPs in relation to human diseases and highlight the most promising synthetic peptides emerging from the clinical trials. Finally, we focused on the limitations and hurdles in terms of cost of production, bioavailability, pharmacokinetic stability and toxicity faced by commercial development and clinical use of the AMPs and strategies to overcome these hurdles.