Antimicrobial peptides, also known as AMPs, are heterogeneous compounds present in various organisms including human beings. Currently, there are hundreds of antimicrobial peptides which have been isolated and properly characterized. They can be thought of as natural microbicides with the ability to selectively cytotoxic bacteria, while at the same time doing minimal harm or damage to the mammalian cells of the host organism.
It has been revealed that this group of peptides has a relatively strong electrostatic attraction to bacterial cells with a net negative charge. Antimicrobial peptides are also known to be able to quickly accumulate at the sites of infections and have minimal cytotoxicity to the host cells. These two factors have made them immensely useful in the formulation of a variety of remedies because of their different mechanisms of action and their specific interactions with the bacterial cells.
In this comprehensive review, we will take a detailed look at the classification of antimicrobial peptides, their design, and their applications. We will also focus on the mechanism of their actions and their selective properties, and describe the various strategies, designs, and potential utilization in the fields of research, medicine, and drug formulation.
Overall overview of antimicrobial peptides
Antimicrobial peptides are currently being viewed as evolutionary biomolecules that play an increasingly important role when it comes to the defense mechanisms of a variety of organisms. They are seen as the organisms at the very first line of defense against pathogenic microbes in higher animals as well as most of the lower forms of life.
For pathogenic and saprophytic microbes, antimicrobial peptides are the only forms of defense for these forms of life. Antimicrobial peptides are known to have selective toxicity –an ability that enables them to attack only pathogenic microbes while not touching the host cell. This is possible because of a fundamental difference in the composition as well as the structure of the host cells when compared to the composition and the structure of the pathogenic bacteria and the yeasts.
Though there are certain antimicrobial peptides that have been discovered to have immunomodulatory effects and chemotactic behaviors, one feature that is common in most of the antimicrobial peptides is the fact that they are amphipathic, but with an overall negative net charge. Presently, it is estimated that there are about 1500 antimicrobial peptides that have been successfully characterized. However, the classification process is still on and this is being done based on the amino acid composition as well as the secondary structures of the peptides. Below is a brief overview of some identifications of certain antimicrobial peptides-:
α-Helical Antimicrobial Peptides
It is estimated that between 30% and 50% of all the antimicrobial peptides identified to this date belong to the α-helical structure. It is believed that the major reason for this classification is the relative ease of these peptides being chemically synthesized, which in turn allows for extensive characterization in the laboratories. Most of the peptides in this category have between 12 and 40 amino acid residues and a plethora of helix stabilizing residues such as lysine and alanine. Whenever they are in aqueous solutions, these peptides tend to be unstructured, though they still tend to retain their helical conformation, especially in the cell membranes. Most of the time, the bulk of these peptides are not strictly α-helical and it is common for them to show signs of internal kink.
β –sheet Antimicrobial peptides
The other major classification of anti-microbial peptides has between two and ten cysteine residues which are responsible for forming between two and five interchain disulfide bonds. Through this bonding interaction, it is possible for the peptides to portray β-sheet conformations. It has been noted that a majority of the β-sheet antimicrobial peptides tend to form part of the defensin family. It has also been noticed that these peptides are evolutionarily conserved across insects, fungi, plants, and vertebrae animals.
Defensins mainly comprise two to three antiparallel β –sheets stabilized by three to four intramolecular disulfide bonds. But in the case of unstructured segments usually found present in the N and the C-termini, this kind of stabilization is never noticed. Unlike the α-helical antimicrobial peptides, which are generally unstructured in aqueous solutions, it has been observed that most of the defensins tend to maintain a very compact and steady globular structure whenever they are in aqueous solutions.
Besides the general similarity in secondary structure, most of the defensins derived from mammals have two additional common features – a bulge caused by conserved glycine and cysteine and a protruding loop that is a result of conserved glutamine/arginine salt bridge.
Antimicrobial Peptides endowed with specific amino acids
Studies have shown that a small number of antimicrobial peptides have high concentrations of certain amino acids, with the commonly noted amino acids being histidine, glycine, tryptophan, and proline. The bulk of the representative members of this minority class of amino acids include tryptophan-rich bovine indolicidin and porcine tritrpticin, histidine-rich human histatin, and proline-rich porcine.
Because of their unusual composition of amino acids, most of these peptides have been found to have highly variable secondary structures. For example, indolicidin, which is known to have 13 amino acids, portrays a largely extended conformation whenever is it in the presence of zwitterionic micelles that comprises substances such as anionic sodium-dodecyl sulfate.
Mechanisms for cell selectivity and specificity of antimicrobial peptides
Major differences in the composition of the microbial and the host cell membranes, as well as the architecture play a major role in the selectivity of antimicrobial peptides. Regulation of expressions or localization of the peptides is also believed to be one of the factors responsible for preventing unwanted interactions with any cells that may vulnerable to the host cells.
Selective Cell Toxicity and Target Specificity
A biological membrane may be simply described as a fluid mosaic containing phospholipids spread in proteins. In most organisms, it is also possible for sterols and glycerides to compose part of the biochemical architecture and the surface topology of the cell membranes.
However, there exist monumental differences between animal cells and microbial cell membranes. It is this fundamental difference that makes it possible for antimicrobial peptides to differentiate between the cells and to have the ability to selectively target one over the other.
The composition, charge, and hydrophobicity of the cell membranes
The major factor of nearly all natural biomembranes is the structure known as the phospholipid bilayer. These layers are usually amphipathic. This simply means that they have both hydrophilic and hydrophobic regions. But, prokaryotic and eukaryotic cell membranes tend to be slightly different when it comes to the specific composition of the cell energetics.
Sphingomyelin, phosphatidylcholine, and phosphatidylethanolamine don’t show any changes in their charges when they are under normal physiological conditions. Sterols such as ergosterol, which are found in plenty in eukaryotic membranes but very rare in prokaryotic membranes, also tend to generally have a neutral net charge.
Hydroxylated phospholipids such as cardiolipin tend to have a net negative charge when under physiological conditions. With this information, it can be clearly seen that the charge of the membrane changes as a result of both the ratio as well as the location of the various phospholipids.
What you need to know about membrane asymmetry
It should be noted that cellular membranes are neither static nor symmetric, but the differences that exist between the mammalian and microbial phospholipid bilayers make it possible for antimicrobial peptides to accurately mark out their targets. In certain cells like those of bovine erythrocyte, it is estimated that less than 3% of the total PE content is found on the outer membrane leaflet.
The differences that exist between membrane saturation, symmetry, and the composition stoichiometry of the phospholipid bilayers have a major impact on the membrane’s fluidity as well as a phase transition. Additionally, it is also possible that the net charge of the outer and the inner leaflets of the cellular bilayer membranes may also be different.
Microbial ligands and receptors as targets for antimicrobial peptides
Several scientific studies have shown that the L and D-amino acid versions of antimicrobial peptides may portray similar binding affinities to those of the target cells. This subtly suggests that there may be certain stereotypic receptors that may not have a role to play when it comes to targeting the pathogenic cells.
However, there are also several studies that seem to refute this notion and instead attempt to suggest that there are certain proteins present in the microbial cell membrane that may serve as binding targets for certain classes of antimicrobial peptides such as histatins. With such, there could be more credence to the findings that histatines are usually involved with the local defense mechanisms targeting particular pathogens.
Some studies suggest that anionic components of the cell membranes may be used as pseudoreceptors which are responsible for making the initial interaction between the microbial cell target and the antimicrobial peptide. Consequently, antimicrobial binding receptors are currently being seen as an alternative pathway for the interaction of antimicrobial peptides with the bacterial cell envelope.
The transmembrane potential of antimicrobial peptides
The transmembrane potential of antimicrobial peptides is another mechanism through which the antimicrobial peptides and mammalian cells vary sharply. The major difference between the two is the charge separation that is present between the outer layers and inner layers of the cytoplasmic membranes. Transmembrane potential refers to an electrochemical gradient created by the differing rates or proton exchange that normally take place across the cell membranes. In a normal mammalian cell, the transmembrane potential is between -90 and -110mV.
In pathogenic bacteria, however, the transmembrane potential is normally observed to be anywhere between -130 and -150mV. This is a significant difference in the electrochemical potential and it is believed to be another factor that may be responsible for antimicrobial peptides’ ability to distinguish between the target cells and the host cells.
Selective Toxicity Based on Antimicrobial Peptide Design
Most of the antimicrobial peptides tend to have extended or unstructured conformations whenever they are in the aqueous intracellular environment. However, in the presence of intramolecular bonds, this doesn’t seem to be the case since specific conformations are synonymous to varying environments as a result of induced rigidity.
After the antimicrobial peptide is strongly bound to the cell membrane of the pathogenic microbe, there are very good chances that it may experience very heavy conformational changes which may lead to them developing very specific conformations such as the α-helix structure. There are several studies that tend to suggest that the inherent or dynamic conformation of the antimicrobial peptides may have a huge impact when it comes to their selective cytotoxicity. Also, several studies suggest that antimicrobial peptides may have to undergo various conformational transitions, oligomerization, and self-association within the larger pathogen membrane.
However, it is not possible for the host cell membrane to increase cell-specific toxicity. In a study conducted by Zhang and coworkers where synthetic test peptides which were uniformly cationic were used with varied conformation and featured extended cyclic structures, it was observed that all the test peptides had the ability to interact and penetrate lipid monolayers which were made of a negatively charged phospholipid layer.
However, the same study determined that only peptides with helical and extended structures had the ability to interact with the more neutrally charged membranes. In the same study, it was also observed that peptides with β-sheet structures could translocate phospholipids from the inner leaflet to the outer leaflet when the concentration of the medium was lower than the concentrations required to permeate the membrane.
In yet another study by Kol and coworkers, it was observed that peptides with comparable conformation, but which had a rich concentration of lysine and histidine and had a low concentration of tryptophan, may also affect the significant levels of phospholipid translocation. From these studies, it can be easily concluded that antimicrobial peptides have the ability to interact with phospholipid membranes that have very specific compositions and symmetry. However, they don’t have the ability to affect the modeling of the membranes in certain specific cells.
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