Cationic Antimicrobial Peptides: Definition, Structure, Applications and Examples

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What are the cationic antimicrobial peptides?

Cationic antimicrobial peptides (CAMPs) are peptides recognized for their capacity to target and damage microbial cell membranes. These peptides are often short, amphipathic molecules containing both hydrophobic and cationic sections, allowing them to engage with negatively charged elements on microbial surfaces. CAMPs are a component of the innate immune response in several species, including humans, plants, and animals, and exhibit efficacy against a wide range of pathogens, encompassing bacteria, fungi, viruses, and some parasites. CAMP genes undergo positive selection, and CAMPs represent one of the most quickly changing categories of mammalian proteins, exhibiting significant variation even within primate species. Certain CAMPs are preserved throughout different mammalian lineages, whereas others appear to have emerged, vanished, or proliferated through gene duplication in certain mammalian families.

Biological function of cationic antimicrobial peptides. (Zhang L., et al., 2016)

Structure of cationic antimicrobial peptides

The short size (about 12-50 amino acids long), hydrophobicity, and cat-ionicity (with an overall charge range from +2 to +9) are commonalities across all naturally occurring cationic antimicrobial peptides. As a component of their innate immune responses, all live eukaryotic and prokaryotic organisms manufacture these peptides, which are typically encoded by genes and expressed either constitutively or inducibly (in response to signals received from infectious or inflammatory agents). Even within a same structural family, there is a lack of sequence homology among these peptides, which may explain why they have persisted through evolution. There are typically four categories of cationic anti-microbial peptides, including synthetic ones: a-helical, β-sheet, extended structures rich in certain amino acids, and loop peptides. The two most prevalent categories are a-helical and β-sheet, respectively. Although there is variation in their primary and secondary structures, when they engage with a membrane, they produce amphiphilic molecules with hydrophobic and charged/polar residues separating into patches, or faces. This similar structural arrangement is maintained at the tertiary level.

Diversity of human cationic antimicrobial peptides. (Peschel A., et al., 2006)

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Examples and mechanisms of cationic antimicrobial peptides

It is often the capacity of CAMPs to bind to bacterial cell walls or membranes that determines their killing potential. In order to bind preferentially to negatively charged bacterial membranes, CAMPs often display a net positive charge and a large ratio of hydrophobic amino acids. A mechanism other than enzymatic degradation is triggered when CAMPs bind to the bacterial cell membrane. The unique makeup of microbial and cellular membranes accounts for species selectivity. There are two mechanisms of CAMPs: membrane-disruptive and membrane-nondisruptive ways.

Membrane-disruptive CAMPs

Once CAMPs bind to phospholipids, interactions with the cytoplasmic membrane can commence. The CAMPs stay bound at the interface of the hydrophilic lipid headgroups and the hydrophobic fatty acyl chains, parallel to the membrane plane, as long as the peptide:lipid ratio is low. The four suggested models—barrel-stave, aggregation, carpet, and toroidal pore—describe how peptides might aggregate and/or reorient inside the membrane when the peptide:lipid ratio rises, therefore disrupting membrane integrity. The aggregate model illustrates how cationic antimicrobial peptides can effect death by both membrane permeability and internal target assault, however no single model is more accurate than the others. Each mechanism has been tested with chosen cationic antimicrobial peptides.

Barrel-stave model: The barrel-stave architecture is occasionally referred to as a helical bundle, as exemplified by the non-cationic peptide alamethicin and gramicidin S. Upon their interaction with the cytoplasmic membrane and reaching a crucial peptide:lipid ratio, the peptides reorient perpendicularly and are hypothesized to traverse the lipid bilayer. Their orientation is such that the hydrophobic side chains are directed towards the hydrophobic core of the membrane, while the polar side chains are posited to face inward, forming a hydrophilic aperture that traverses the membrane's breadth. The formation of the pore may subsequently result in the leaking of cellular contents. Nonetheless, this model fails to elucidate the mechanisms underlying pore formation in most instances, as the channels that develop are frequently irregular in both size and duration, and exhibit mild cation selectivity; conversely, if numerous cationic headgroups were aligned within the lumen of a narrow channel, one would anticipate an anion specificity.

Toroidal pore model: In the toroidal pore model, also referred to as the wormhole model (e.g., magainin 2), CAMPs adhere to the membrane, prompting the lipids to invaginate and create a channel bordered by lipid headgroups and associated peptides at the membrane interface, thereby establishing a continuous conduit between the inner and outer leaflets. The peptides predominantly maintain their association with the lipid headgroups during the process, in contrast to the barrel-stave model, where peptides are first linked to the headgroups before engaging with the lipid tails. The resultant hole causes cellular leakage.

Carpet model: In contrast to previous models, the carpet model (e.g., cecropin) indicates that CAMPs do not penetrate the membrane but instead remain linked with the interfacial region of the outer leaflet of the cytoplasmic membrane. At a pivotal juncture, the accumulation of peptides creates a 'carpet' that can disrupt the bilayer by impairing membrane electrostatics, leading to the membrane's collapse into a micellar structure. Cellular demise would result from the depletion of cytoplasmic constituents. This model predicts a critical concentration at which the membrane would collapse, however this is not consistently supported by experimental evidence.

Aggregate model: The aggregation model resembles the detergent model (e.g., polyphemusin and indolicidin). Upon binding to the membrane interface, at enough concentrations, the peptides reorient, facilitating the formation of micelle-like complexes with the lipids that traverse the lipid bilayer in a peptide-lipid complex. Transmembrane random aggregates of lipids, cationic peptides, and water can create a channel for ion leakage, resulting in cell death due to the loss of cytoplasmic contents, or they may spontaneously disintegrate, facilitating the translocation of cationic peptides into the cytoplasm, where they can induce internal cytotoxicity. This model elucidates the mechanism by which CAMPs can compromise membrane integrity and/or facilitate peptide translocation over the membrane into the cytoplasm.

Membrane-nondisruptive CAMPs

New research indicates that certain CAMPs can target interior organelles, either as their main mode of action after crossing the membrane or as a synergistic effect with (often partial) rupture of the membrane. Cationic antimicrobial peptides are considered "dirty drugs" by many researchers, and their findings point to a number of potential interior targets. By interacting with various anionic targets, such as nucleic acids to influence RNA or DNA synthesis, cellular enzymes, cell division, or membranes, CAMPs are able to exert their multitarget mode of action.

CAMPs that act on nucleic acids: Evidence suggests that several cationic antimicrobial peptides interact with DNA and RNA. Buforin Il, a 21-amino acid LysC endoproteinase derivative of the 39-amino acid buforin I, was isolated from the Asian toad Bufo bufo gargarizans and is one of the peptides that interacts with DNA that has been examined more extensively. There was evidence that buforin II could move across liposome lipid bilayers without causing lipid flip-flop. Similarly, fluoroisothiocyanate (FITC)-labeled buforin II could cross the E. coli cytoplasmic membrane and accumulate inside, even below its minimum inhibitory concentration (MIC). This, along with the fact that it didn't disrupt the membrane or cause cell lysis, suggested that it likely had an internal target. Curiously, buforin forms a disrupted a-helix with a prolinehinge, which is a common motif among antimicrobial peptides. However, when the proline residue was removed, the peptide no longer had the ability to enter cells. Instead, it localized on the cell surface and permeabilized the cell membrane. This finding suggests that even minor structural changes can have a significant impact on the mechanism. The ability of buforin II to bind to DNA was confirmed by gel retardation experiments. Not surprisingly, this is because buforin I shares 37 out of 39 amino acids in homology with the N-terminal region of Xenopus histone H2A, a DNA-binding nuclear protein. Other cationic antimicrobial peptides, like hipposin and parasin I, also share homology with the N-termina region of the histone H2A, and some histones have antibacterial activity as well. However, the mechanisms by which these peptides work are still poorly understood.

CAMPs that act on protein synthesis: One 39-amino acid cecropin, PR39, was discovered to suppress protein synthesis in E. coli after having been extracted from pig intestines and subsequently from pig neutrophils. The breakdown of DNA replication enzymes may have contributed to the end of DNA synthesis together with the end of protein synthesis. Experiments with the indolicidin variation CP10A, which has three proline-to-alanine substitutions, showed that this peptide may disrupt histidine incorporation at double the minimum inhibitory concentration (MIC), hence inhibiting protein synthesis in Staphylococcus aureus. Another pathway that CP10A impacted was the synthesis pathway for macromolecules. Also demonstrated to affect protein synthesis was a pleurocidin derivative derived from winter salmon, which inhibited histidine incorporation in E.coli at the minimum inhibitory concentration (MIC). Dermaseptin, pleurocidin, and human defensin human neutrophil peptide-1 are a few more cationic antimicrobial peptides that have been shown to affect protein synthesis. Please be aware that the production of other macromolecules was also impeded by a few of these peptides.

CAMPs that act on translation/protein folding: Bacterial cells, like eurkaryotic cells, have chaperones that help get proteins and peptides folded correctly. Recently, the target of the insect-produced cationic antimicrobial peptides pyrrhocoricin, drosocin, and apidaecin was determined to be a 70-kDa bacterial chaperone, DnaK, using immunoaffinity purification and mass spectrometry. The results showed that these peptides bound exclusively to E. coli DnaK and had no effect on the human chaperone Hsp70 or GroEL, two additional bacterial chaperones. Subsequent research shown that pyr.rhocoricin and drosocin interfered with DnaK's ATPase activity, likely as a result of these peptides attaching to DnaK and blocking its multihelical lid.

Summary of cationic antimicrobial peptides and their mechanisms of action. (Hale J D F., et al., 2007)

Applications of cationic antimicrobial peptides

Antibiotic Alternatives

In light of the alarming increase in antibiotic resistance, CAMPs are being investigated as potential substitutes for traditional antibiotics. They provide hope for the fight against antibiotic-resistant illnesses since they work against germs that are resistant to many drugs.

Infection treatments

Creams, wound dressings, and inhalable medications for respiratory diseases are among possible uses for CAMPs. Because their topical administration reduces systemic toxicity, they are especially useful in treating infections of the skin and wounds, including as burn wounds and diabetic ulcers.

Food preservation

Because of their antimicrobial properties, CAMPs are being studied as a potential natural food preservative that can help keep food fresh and reduce the risk of food poisoning without the need of artificial ingredients.

Cancer treatment

Traditional cancer treatments like chemotherapy have drawbacks including harmful side effects and cancer cells that are able to acquire resistance to many drugs. CAMPs shows promise in overcoming the limitations of traditional chemotherapy. This is due to the fact that some CAMPs show selective cytotoxicity against a wide variety of human cancer cells, including neoplastic cells that have developed a multi-drug resistance. Most CAMPs kill tumor cells by destroying their cell membranes, but some can also cause cancer cells to die by damaging their mitochondria. Additionally, certain CAMPs are very effective at blocking the process of angiogenesis, the formation of new blood vessels, which is linked to the advancement of tumors.

References

1. Zhang L., et al., Antimicrobial peptides, Current biology, 2016, 26(1): R14-R19.
2. Hale J D F., et al., Alternative mechanisms of action of cationic antimicrobial peptides on bacteria, Expert review of anti-infective therapy, 2007, 5(6): 951-959.
3. Peschel A., et al., The co-evolution of host cationic antimicrobial peptides and microbial resistance, Nature Reviews Microbiology, 2006, 4(7): 529-536.
4. Geitani R., et al., Expression and roles of antimicrobial peptides in innate defense of airway mucosa: potential implication in cystic fibrosis, Frontiers in immunology, 2020, 11: 1198.
5. Mader J S., et al., Cationic antimicrobial peptides as novel cytotoxic agents for cancer treatment, Expert opinion on investigational drugs, 2006, 15(8): 933-946.