Immunological Mechanism and Application Prospect of Antibacterial Peptides

Over the past few decades, more than 700 antimicrobial peptides have been discovered. These peptides are classified into two categories, one being non-ribosome synthetic antibacterial peptides such as gramicidin, colistin, bacitracin, and sugar polypeptides, which are mainly produced by bacteria and obtained through structural modifications; The other is a natural antibacterial peptide synthesized from ribosomes, which is a kind of antimicrobial and malignant-cell-toxic peptides produced by the organism during defense reaction against pathogenic microorganisms. These peptides are a class of antibacterial molecules that are encoded by genes and produced by host cells. They are the first immunologically active molecules produced by organisms to adapt to the environment and survive in the course of evolution. Their molecular weight is relatively small, generally around 4000Dr. Around tens of amino acid residues, it is considered as an important medium for innate immunity, for G (upper right +), G (upper right -), mold, spirochetes, viruses (such as influenza virus, herpes virus, and AIDS virus) ), etc. have a strong killing activity. I. Antimicrobial peptides synthesized by non-ribosomes Biosynthesis. Antimicrobial peptides synthesized by non-ribosomes are peptide substances with antibacterial activity secreted by bacteria, fungi, Streptomyces and the like. Studies have shown that these antibacterial peptides are synthesized under the action of peptide synthetase according to the multi-carrier sulfur template mechanism. Peptide synthase genes (such as Bacitracin S biosynthetic opers grsB) are 13kb in size and contain four to six templates. Each template recognizes, activates one amino acid residue, and modifies it when necessary. Add to the peptide chain being synthesized. Among them, the smallest template can activate one amino acid residue or hydroxy acid residue, and the activated amino acid is polymerized to the peptide under the synergistic action of a covalent bond (4′-phosphopantethein) with the template. On the chain and can also stabilize activated amino acid residues. Many peptides containing hydroxyl, L-amino acid and D-acid can be synthesized under the control of this mechanism, and further chemical modifications such as N-based, acylation, glycosylation or heterocyclic formation become Biologically active antimicrobial peptides. 2. Biological activity and antibacterial mechanism. Polymyxin B carries 5 positive charges and Gramicidin S carries 2 positive charges. In essence they are all cationic antimicrobial peptides. Polymyxin has strong antibacterial activity against G (upper right -) bacteria, and the main susceptible strains are Escherichia coli, Salmonella, Pasteurella, Brucella, Vibrio, Shigella, Pseudomonas, etc. Bacillus has a strong bactericidal effect. Gramicidin S and bacitracin mainly act on G (upper right +) bacteria. Bacitracin is effective against resistant Staphylococcus aureus, Enterococci, and Streptococcus and it is also effective against spirochetes and actinomycetes, but it is not effective against G (upper right-) bacteria. According to data, Gramicidin S is highly active against G (upper right -) bacteria and Candida albicans. Bacitracin inhibits bacterial growth by inhibiting the conversion of G (preferred +) peptidoglycan precursors to pyrophosphate sterols; polymyxins kill bacteria by disrupting bacterial membranes. The role of; other non-ribosomal synthesis of antibacterial peptides, such as streptogramin by inhibiting bacterial protein synthesis play a bacteriostatic effect. 3. Clinical application. Colistin is a methylthio derivative of cationic lipopeptide polymyxin E. The use of aerosol for the treatment of pulmonary P. aeruginosa infection has been successful. The purpose of chemical modification of natural lipopeptides is to reduce toxicity in systemic applications. Part of the toxicity of natural lipopeptides may come from the tail of the peptide chain. Some scholars believe that the non-acylated derivatives of polymyxin will have a strong toxic reaction when used systemically. Non-acyl Gramicidin S can cause Hemolysis. Therefore, the widespread use of this class of antibiotics has been limited. 4. Application prospects. The discovery of non-ribosome synthetic antibacterial peptides provides humans with a wealth of new antibacterial drug resources. At present, people develop such antibiotics mainly through 3 ways. First, new antibiotics are sought for the existing antimicrobial peptides and their derivatives through chemical modification. For example, streptavidin is a family of cyclic antimicrobial peptides discovered in the 1950s. Its antibacterial properties are very strong, but its solubility is extremely poor. Two kinds of water-soluble semi-synthetic streptogramins, dalfopristin, have been obtained by chemical means. Quinupristin, a Phase III clinical trial has been completed as a anti-drug resistant G (upper right +) strain. Second, using templates to synthesize antibacterial peptides is an exciting method. Schneider et al. pointed out that the new peptide synthesis template was assembled to obtain a new structure. In this way, it is possible to obtain a large number of antimicrobial peptide backbone chemical structures. Third, chemical synthesis is performed for the precursor using the resulting chemical structure. Gramicidin is a good example. Gramicidin S derivatives with different sizes of parent rings, different charge, amino acid sequence and hydrophobicity are highly selective to bacteria. 2. Antimicrobial peptides synthesized by ribosomes As early as a few centuries ago, people knew that medicinal properties of frog skin were available, but it was not until 1962 that Kiss and Michl discovered antibacterial and hemolytic effects from the skin secretions of Bombina variegata. The peptides, and ultimately the 22-amino acid peptide (bombinin) were isolated from it. It was only known that the antibacterial effect of the skin was due to the presence of active substances such as antimicrobial peptides. In 1972, the antibacterial and hemolytic peptide melittin was isolated from bee venom. Since then, the structure and mechanism of action of such cationic antimicrobial peptides have been studied in depth. Up to now, more than 700 antimicrobial peptides have been found in various organisms. 1. distributed. All organisms including bacteria, fungi, insects, tunicates, amphibians, crustaceans, birds, fish, mammals (including humans) and plants can produce antimicrobial peptides. In mammals, antimicrobial peptides (such as defensins) are the major protein component of neutrophils (10% to 18% of total protein). Increased concentration of antimicrobial peptides on damaged mucosal surfaces becomes an important material for mucosal defense. 2. Biological activity and antibacterial mechanism. Antimicrobial peptides are active against G (upper right +) bacteria, G (upper right -) bacteria, fungi, enveloped viruses, cancer cells and parasites, can promote wound healing and also have chemotaxis. A good antimicrobial peptide should be effective against most bacteria and is highly effective against multi-resistant P. aeruginosa, meticillin-resistant S. aureus, and Stenotrophomonas maltophilia. Some bacteria also develop resistance to antibacterial peptides but are only mildly resistant. Although antimicrobial peptides vary widely in length, almost all antimicrobial peptides are cationic in nature, and their higher structures appear as either а-helix or β-sheet, and amphiphilic structures are their common features. Boman et al. found that porcine cecropin P1 was bactericidal by lysing bacteria, while PR-39 was used for bactericidal purposes by blocking bacterial protein and DNA synthesis. According to Christensen et al., at the interface between the plasma membrane and the aqueous phase, the antibacterial peptide and the lipid bilayer first approach each other by electrostatic attraction. With the help of the flexibility of the connection structure between the N- and C-termini in the molecule, the antimicrobial peptide molecule is The hydrophobic end of the membrane is inserted into the plasma membrane, and then the amphipathic а-helix is ​​also inserted into the plasma membrane. The defensins isolated from neutrophils have an amphiphilic β-sheet configuration and the bactericidal activity lies in infiltration of the outer and inner membranes of E. coli, similar to the formation of ion channels on the membrane by cecropins. Whether it is an antibacterial peptide having an а-helical or β-sheet structure, it has the ability to form channels, ie, channel formation plays an important role in antibacterial activity. In addition, cation characteristics are also essential for antibacterial activity. For G (upper right -) bacteria, antimicrobial peptides interact with negatively charged lipopolysaccharides on their adventitia, disrupting the outer membrane structure to cross the inner membrane, while G (upper right +) bacteria do not have lipopolysaccharide membranes, but The surface is negatively charged due to the existence of truncated acid, uronic acid, teichoic acid, and amino acid carboxyl groups in peptidoglycan, and the antimicrobial peptide can also disrupt the peptidoglycan layer and penetrate the plasma membrane. Little is known about the antifungal mechanism of antimicrobial peptides. Some people think that antibacterial peptides can change the morphology of fungi, the rapid outflow of internal ions, or inhibition of mitochondrial synthesis energy. Some antimicrobial peptides are active against trypanosomes, malaria and nematodes. Antimicrobial peptides have also been reported as anti-cancer cells, but they are also likely to be toxic to normal cells. 3. The role of antimicrobial peptides in innate immunity. Cationic antimicrobial peptides play a very important role in the host's natural immunity. Variations in the regulatory genes or signal genes of Drosophila can affect the expression of many peptides, leading to increased susceptibility to bacteria and fungi. Wilson et al. isolated an enzyme matrilysin from mice to convert intestinal defensins into active defensins. Removing the enzyme gene can block the conversion of intestinal defensins into active defensins, resulting in a 10-fold increase in the susceptibility of mice to microorganisms. Lack of а-defensins in patients with specific particle deficiency syndrome can often result in serious bacterial infections. It can be seen that antimicrobial peptides play a very important role in host anti-infection. Antimicrobial peptides neutralize bacterial signal molecules such as G (upper right -) bacterial lipopolysaccharides, G (upper right +) mycelial teichoic acids, and bacterial unmethylated CpG DNA. These signaling molecules can bind to Toll-like receptors on the host cell surface, initiating signal cascade amplification and positive regulation of cytokines (eg, TNF, IL-6, etc.). Low concentrations of bacterial signaling molecules can trigger beneficial inflammatory reactions and fever in the body, but if the reaction is severe or prolonged, it can lead to systemic circulatory disorders, organ failure, and even death. Antimicrobial peptides can inhibit the binding of lipopolysaccharide and serum lipopolysaccharide binding protein factors and prevent endotoxemia and death. Antimicrobial peptides are also involved in other reactions in the host's natural immunity, such as stimulating the chemotaxis of monocytes and neutrophils, promoting mast cell histamine release, inhibiting cathepsins, and promoting wound healing. Microassay experiments have demonstrated that antimicrobial peptides can selectively regulate the expression of more than 30 genes. However, there is still no complete animal model data to prove that antimicrobial peptides play an important role in these reactions. 4. Clinical application and existing problems. At present, there are only 5 clinical trials on antimicrobial peptides. (1) IB-367, an integrin-like molecule, is a Phase III clinical trial to treat multi-microbial involvement of oral mucosal inflammation caused by radiotherapy and chemotherapy in cancer patients; (2) IB-367 aerosol treatment of cystic cellulose Phase II clinical trial of pulmonary Pseudomonas infection in patients with pneumoconiosis; (3) Phase III clinical trials of Indolicidin to prevent infection in central venous cannula sites; (4) Phase III clinical trials of MBI-226 to prevent infection in central venous cannula sites Test; (5) Indolicidin-like peptides for the treatment of acute acne in Phase II clinical trials. Before any new drug can be successfully applied to the clinic, it must undergo a series of tests. These tests can prove that the drug has good activity, stable formulation, suitable production method, sufficient stability in the body and very low The toxicity. In theory, antibacterial peptides should be ideal candidates for antibacterial drugs, but there is almost no complete animal in vivo experimental data. Zhang et al. found that polyphemusin I isolated from horseshoe crabs has good antibacterial activity in vitro, but animal model experiments have shown little resistance to infection. After modified by polyphemusin I, the antibacterial activity in vitro decreased, and the anti-infective capacity in vivo increased. The main reason for the lack of in vivo antibacterial activity of antimicrobial peptides may be the decrease of the stability of antimicrobial peptides caused by host protease. This shortcoming can be overcome by improving the formulation (such as masking antimicrobial peptides with liposomes), using prodrugs, binding proteolytic enzyme-binding peptides, or performing sequence modifications. Varbanac et al. pointed out that the antibacterial peptide precursor may be a protein inhibitor, and the antibacterial peptide may be converted from a protease substrate into a protease inhibitor through appropriate sequence modification.