Inhibitory and anti‐inflammatory effects of two antimicrobial peptides moronecidin and temporin‐1Dra against Propionibacterium acnes in vitro and in vivo

Proliferation of Propionibacterium acnes (P. acnes) is one of the main pathogenetic mechanisms of acne. Antimicrobial peptides with low‐drug resistance and nonresidual are potential anti‐acne agents. In this study, two antimicrobial peptides named temporin‐1Dra and moronecidin were synthesized and tested their antimicrobial activity against P. acnes in vitro and in vivo. These two peptides inhibited the growth of Escherichia coli, Staphylococcus aureus, Candida albicans, and P. acnes. The minimal inhibitory concentrations (MICs) of temporin‐1Dra and moronecidin to P. acnes were 30 and 10 μM, respectively. Both peptides exhibited strong resistance to heat and pH, but no obvious cytotoxicity to HaCaT cells. They also displayed persistent antimicrobial activities in the microbial challenge test. In the P. acnes‐induced inflammation mouse model, moronecidin significantly decreased the ear swelling thickness in a concentration‐dependent manner. At the 14th day after injection, 20 μg/day moronecidin reduced the ear swelling thickness to 46.15 ± 5.23% compared with the normal cream group. Tissue staining showed that moronecidin effectively reduced abscess and thickness of the dermis layer. Our results indicate that the antimicrobial peptide moronecidin could be developed as a potential natural anti‐acne agent in the cosmetics or pharmaceutical industries.     

A novel cysteine‐free venom peptide with strong antimicrobial activity against antibiotics‐resistant pathogens from the scorpion Opistophthalmus glabrifrons

Antibiotic‐resistant bacteria, such as methicillin‐resistant Staphylococcus aureus and vancomycin‐resistant Enterococcus, pose serious threat to human health. The outbreak of antibiotic‐resistant pathogens in recent years emphasizes once again the urgent need for the development of new antimicrobial agents. Here, we discovered a novel antimicrobial peptide from the scorpion Opistophthalmus glabrifrons, which was referred to as Opisin. Opisin consists of 19 amino acid residues without disulfide bridges. It is a cationic, amphipathic, and α‐helical molecule. Protein sequence homology search revealed that Opisin shares 42.1–5.3% sequence identities to the 17/18‐mer antimicrobial peptides from scorpions. Antimicrobial assay showed that Opisin is able to potently inhibit the growth of the tested Gram‐positive bacteria with the minimal inhibitory concentration (MIC) values of 4.0–10.0 μM; in contrast, it possesses much lower activity against the tested Gram‐negative bacteria and a fungus. It is interesting to see that Opisin is able to strongly inhibit the growth of methicillin‐ and vancomycin‐resistant pathogens with the MICs ranging from 2.0 to 4.0 μM and from 4.0 to 6.0 μM, respectively. We found that at a concentration of 5 × MIC, Opisin completely killed all the cultured methicillin‐resistant Staphylococcus aureus. These results suggest that Opisin is a promising therapeutic candidate for the treatment of the antibiotic‐resistant bacterial infections. Copyright © 2015 European Peptide Society and John Wiley & Sons, Ltd.     

Synthesis of poly(3‐hydroxybutyrate‐co‐4‐hydroxybutyrate)/chitosan/silver nanocomposite material with enhanced antimicrobial activity

This work aims to shed light in the fabrication of poly(3‐hydroxybutyrate‐co‐44%‐4‐hydroxybutyrate)[P(3HB‐co‐44%4HB)]/chitosan‐based silver nanocomposite material using different contents of silver nanoparticle (SNP); 1–9 wt%. Two approaches were applied in the fabrication; namely solvent casting and chemical crosslinking via glutaraldehyde (GA). A detailed characterization was conducted in order to yield information regarding the nanocomposite material. X‐ray diffraction analysis exhibited the nature of the three components that exist in the nanocomposite films: P(3HB‐co‐4HB), chitosan, and SNP. In term of mechanical properties, tensile strength, and elongation at break were significantly improved up to 125% and 22%, respectively with the impregnation of the SNP. The melting temperature of the nanocomposite materials was increased whereas their thermal stability was slightly changed. Scanning electron microscopy images revealed that incorporation of 9 wt% of SNP caused agglomeration but the surface roughness of the material was significantly improved with the loading. Staphylococcus aureus and Escherichia coli were completely inhibited by the nanocomposite films with 7 and 9 wt% of SNP, respectively. On the other hand, degradation of the nanocomposite materials outweighed the degradation of the pure copolymer. These bioactive and biodegradable materials stand a good chance to serve the vast need of biomedical applications namely management and care of wound as wound dressing. © 2014 American Institute of Chemical Engineers Biotechnol. Prog., 30:1469–1479, 2014     

High‐yield recombinant expression of the chicken antimicrobial peptide fowlicidin‐2 in Escherichia coli

The antimicrobial peptide fowlicidin‐2 identified in chicken is a member of the cathelicidins family. The mature fowlicidin‐2 possesses high antibacterial efficacy and lipopolysaccharide (LPS) neutralizing activity, and also represents an excellent candidate as an antimicrobial agent. In the present study, the recombinant fowlicidin‐2 was successfully produced by Escherichia coli (E. coli) recombinant expression system. The gene encoding fowlicidin‐2 with the codon preference of E. coli was designed through codon optimization and synthesized in vitro. The gene was then ligated into the plasmid pET‐32a(+), which features fusion protein thioredoxin at the N‐terminal. The recombinant plasmid was transformed into E. coli BL21(DE3) and cultured in Luria‐Bertani (LB) medium. After isopropyl‐β‐D‐thiogalactopyranoside (IPTG) induction, the fowlicidin‐2 fusion protein was successfully expressed as inclusion bodies. The inclusion bodies were dissolved and successfully released the peptide in 70% formic acid solution containing cyanogen bromide (CNBr) in a single step. After purification by reverse‐phase high‐performance liquid chromatography (RP‐HPLC), ～6.0 mg of fowlicidin‐2 with purity more than 97% was obtained from 1 litre of bacteria culture. The recombinant peptide exhibited high antibacterial activity against the Gram‐positive and Gram‐negative bacteria, and even drug‐resistant strains. This system could be used to rapidly and efficiently produce milligram quantities of a battery of recombinant antimicrobial peptides as well as for large‐scale production. © 2015 American Institute of Chemical Engineers Biotechnol. Prog., 31:369–374, 2015     

Effects of dimerization of the cell‐penetrating peptide Tat analog on antimicrobial activity and mechanism of bactericidal action

The cell‐penetrating peptide Tat (48–60) (GRKKRRQRRRPPQ) derived from HIV‐1 Tat protein showed potent antibacterial activity (MIC: 2–8 µM ). To investigate the effect of dimerization of Tat (48–60) analog, [Tat(W): GRKKRRQRRRPWQ‐NH₂], on antimicrobial activity and mechanism of bactericidal action, its dimeric peptides, di‐Tat(W)‐C and di‐Tat(W)‐K, were synthesized by a disulfide bond linkage and lysine linkage of monomeric Tat(W), respectively. From the viewpoint of a weight basis and the monomer concentration, these dimeric peptides displayed almost similar antimicrobial activity against six bacterial strains tested but acted more rapidly against Staphylococcus aureus on kinetics of bactericidal activity, compared with monomeric Tat(W). Unlike monomeric Tat(W), these dimeric peptides significantly depolarized the cytoplasmic membrane of intact S. aureus cells at MIC and induced dye leakage from bacterial‐membrane‐mimicking egg yolk L ‐α‐phosphatidylethanolamine/egg yolk L ‐α‐phosphatidyl‐DL ‐glycerol (7:3, w/w) vesicles. Furthermore, these dimeric peptides were less effective to translocate across lipid bilayers than monomeric Tat(W). These results indicated that the dimerization of Tat analog induces a partial change in the mode of its bactericidal action from intracellular target mechanism to membrane‐targeting mechanism. Collectively, our designed dimeric Tat peptides with high antimicrobial activity and rapid bactericidal activity appear to be excellent candidates for future development as novel antimicrobial agents. Copyright © 2009 European Peptide Society and John Wiley & Sons, Ltd.     

Immobilization and orientation‐dependent activity of a naturally occurring antimicrobial peptide

A naturally occurring antimicrobial peptide, SMAP‐29, was synthesized with an n‐terminal or c‐terminal cysteine, termed c_SMAP and SMAP_c, respectively, for site‐directed immobilization to superparamagnetic beads. Immobilized SMAP orientation‐dependent activity was probed against multiple bacteria of clinical interest including Acinetobacter baumannii, Pseudomonas aeruginosa, Bacillus anthracis sterne and Staphylococcus aureus. A kinetic microplate assay was employed to reveal both concentration and time‐dependent activity for elucidation of minimum bactericidal concentration (MBC) and sub‐lethal effects. Immobilized SMAP activity was equivalent or reduced compared with soluble SMAP_c and c_SMAP regardless of immobilization orientation, with only one exception. A comparison of immobilized SMAP_c and c_SMAP activity revealed a bacteria‐specific potency dependent on immobilization orientation, which was contrary to that seen in solution, wherein SMAP_c was more potent against all bacteria than c_SMAP. Sub‐MBC kinetic studies displayed the influence of peptide exposure to the cells with multiple bacteria exhibiting increased susceptibility and efficacy at lower concentrations upon extended exposure (i.e. MBC enhancement). For instances in which complete killing was not achieved, two predominant effects were evident: retardation of growth rate and an increased lag phase. Both effects, seen independently and concomitantly, indicate some degree of induced cellular damage that can serve as a predictor toward eventual cell death. SMAP_c immobilized on glass through standard silanization chemistry was also investigated to ascertain the influence of substrate on activity against select bacteria. Published 2015. This article is a U.S. Government work and is in the public domain in the USA.     

Enhancement of the anti‐inflammatory activity of temporin‐1Tl‐derived antimicrobial peptides by tryptophan,arginine and lysine substitutions

Temporin‐1Tl (TL) is a 13‐residue frog antimicrobial peptide (AMP) exhibiting potent antimicrobial and anti‐inflammatory activity. To develop novel AMP with improved anti‐inflammatory activity and antimicrobial selectivity, we designed and synthesized a series of TL analogs by substituting Trp, Arg and Lys at selected positions. Except for Escherichia coli and Staphylococcus epidermidis, all TL analogs exhibited retained or increased antimicrobial activity against seven bacterial strains including three methicillin‐resistant Staphylococcus aureus strains compared with TL. TL‐1 and TL‐4 showed a little increase in antimicrobial selectivity, while TL‐2 and TL‐3 displayed slightly decreased antimicrobial selectivity because of their about twofold increased hemolytic activity. All TL analogs demonstrated greatly increased anti‐inflammatory activity, evident by their higher inhibition of the production tumor necrosis factor‐α (TNF‐α) and nitric oxide and the mRNA expression of inducible nitric oxide synthase and TNF‐α in lipopolysaccharide (LPS)‐stimulated RAW264.7 macrophage cells, compared with TL. Taken together, the peptide anti‐inflammatory activity is as follows: TL‐2 ≈ TL‐3 ≈ TL‐4 > TL‐1 > TL. In addition, LPS binding ability of the peptides corresponded with their anti‐inflammatory activity. These results apparently suggest that the anti‐inflammatory activity of TL analogs is associated with the direct binding ability between these peptides and LPS. Collectively, our designed TL analogs possess improved anti‐inflammatory activity and retain antimicrobial activity without a significant increase in hemolysis. Therefore, it is evident that our TL analogs constitute promising candidates for the development of peptide therapeutics for gram‐negative bacterial infection. Copyright © 2015 European Peptide Society and John Wiley & Sons, Ltd.     

Short KR‐12 analogs designed from human cathelicidin LL‐37 possessing both antimicrobial and antiendotoxic activities without mammalian cell toxicity

KR‐12 (residues 18–29 of LL‐37) was known to be the smallest peptide of human cathelicidin LL‐37 possessing antimicrobial activity. In order to optimize α‐helical short antimicrobial peptides having both antimicrobial and antiendotoxic activities without mammalian cell toxicity, we designed and synthesized a series of KR‐12 analogs. Highest hydrophobic analogs KR‐12‐a5 and KR‐12‐a6 displayed greater inhibition of lipopolysaccharide (LPS)‐stimulated tumor necrosis factor‐α production and higher LPS‐binding activity. We have observed that antimicrobial activity is independent of charge, but LPS neutralization requires a balance of hydrophobicity and net positive charge. Among KR‐12 analogs, KR‐12‐a2, KR‐12‐a3 and KR‐12‐a4 showed much higher cell specificity for bacteria over erythrocytes and retained antiendotoxic activity, relative to parental LL‐37. KR‐12‐a5 displayed the strongest antiendotoxic activity but almost similar cell specificity as compared with LL‐37. Also, these KR‐12 analogs (KR‐12‐a2, KR‐12‐a3, KR‐12‐a4 and KR‐12‐a5) exhibited potent antimicrobial activity (minimal inhibitory concentration: 4 μM) against methicillin‐resistant Staphylococcus aureus. Taken together, these KR‐12 analogs have the potential for future development as a novel class of antimicrobial and anti‐inflammatory therapeutic agents. Copyright © 2013 European Peptide Society and John Wiley & Sons, Ltd.     

Effective antimicrobial activity of Cbf‐K16 and Cbf‐A7A13 against NDM‐1‐carrying Escherichia coli by DNA binding after penetrating the cytoplasmic membrane in vitro

New Delhi metallo‐beta‐lactamase‐1(NDM‐1)‐carrying isolates, which are resistant to most clinical used antibiotics except for tigecycline and colistin, have been found worldwide. Cathelicidin‐BF (BF‐30) is found in the venom of the snake Bungarus fasciatus and exhibits broad antimicrobial activity. Cbf‐K₁₆ and Cbf‐A₇A₁₃ were obtained by mutating Lys¹⁶, Ala⁷, and Ala¹³ of BF‐30, respectively. To investigate their antimicrobial activities against NDM‐1 carrying bacteria, recombinant Escherichia coli BL21 (DE3)‐NDM‐1 with high NDM‐1 activity was constructed by inserting the Klebsiella pneumoniae NDM‐1 gene (GenBank accession no. HQ328085) into a pET28a vector and transforming it into E. coli BL21 (DE3). The peptides showed effective antimicrobial activities against NDM‐1‐carrying E. coli, and the minimum inhibitory concentrations of Cbf‐K₁₆ and Cbf‐A₇A₁₃ were only 4 and 8 µg/ml, whereas those of minimum bactericidal concentrations were 8 and 16 µg/ml, respectively. A time course experiment showed that colony forming unit counts rapidly decreased, and bacteria were thoroughly eliminated within 3 and 6 h by the Cbf‐K₁₆ and Cbf‐A₇A₁₃ treatments, respectively. The peptides penetrated the bacterial cell membrane and enabled β‐galactosidase leakage, and caused the cytoplasmic membrane to become permeable, and finally bound to the DNA. The genomic DNA of E. coli was completely unable to migrate on an agarose gel after Cbf‐K₁₆ treatment (8 µg/ml). These data demonstrated that Cbf‐K₁₆ and Cbf‐A₇A₁₃ possess effective antimicrobial activity against drug‐resistant strains, including NDM‐1 carrying E. coli BL21 (DE3)‐NDM‐1, by binding to DNA after penetrating the cytoplasmic membrane in vitro, which may have potential therapeutic value for the treatment of NDM‐1‐carrying bacterial infections. Copyright © 2013 European Peptide Society and John Wiley & Sons, Ltd.     

High‐molecular‐weight poly(Gly‐Val‐Gly‐Val‐Pro) synthesis through microwave irradiation

In this study, we synthesized a polypeptide from its pentapeptide unit using microwave irradiation. Effective methods for polypeptide synthesis from unit peptides have not been reported. Here, we used a key elastin peptide, H‐GlyValGlyValPro‐OH (GVGVP), as the monomer peptide. It is difficult to obtain poly(Gly‐Val‐Gly‐Val‐Pro) (poly(GVGVP)) from the pentapeptide unit of elastin, GVGVP, via polycondensation. Poly(GVGVP) prepared from genetically recombinant Escherichia coli is a well‐known temperature‐sensitive polypeptide, and this temperature sensitivity is known as the lower critical solution temperature. When microwave irradiation was performed in the presence of various additives, the pentapeptide (GVGVP) polycondensation reaction proceeded smoothly, resulting in a product with a high molecular weight in a relatively good yield. The reaction conditions, like microwave irradiation, coupling agents, and solvents, were optimized to increase the reaction efficiency. The product exhibited a molecular weight greater than Mr 7000. Further, the product could be synthesized on a gram scale. The synthesized polypeptide exhibited a temperature sensitivity that was similar to that of poly(GVGVP) prepared from genetically recombinant E. coli. Therefore, this technique offers a facile and quick approach to prepare polypeptides in large amounts. Copyright © 2016 European Peptide Society and John Wiley & Sons, Ltd.     

The chitin‐binding capability of Cy‐AMP1 from cycad is essential to antifungal activity

Antimicrobial peptides are important components of the host innate immune responses by exerting broad‐spectrum microbicidal activity against pathogenic microbes. Cy‐AMP1 found in the cycad (Cycas revoluta) seeds has chitin‐binding ability, and the chitin‐binding domain was conserved in knottin‐type and hevein‐type antimicrobial peptides. The recombinant Cy‐AMP1 was expressed in Escherichia coli and purified to study the role of chitin‐binding domain. The mutants of Cy‐AMP1 lost chitin‐binding ability completely, and its antifungal activity was markedly decreased in comparison with native Cy‐AMP1. However, the antimicrobial activities of the mutant peptides are nearly identical to that of native one. It was suggested that the chitin‐binding domain plays an essential role in antifungal, but not antimicrobial, activity of Cy‐AMP1. Copyright © 2009 European Peptide Society and John Wiley & Sons, Ltd.     

基于厚壳贻贝Mytilin-1的抗菌肽设计、固相合成及抗菌谱分析

贻贝抗菌肽Mytilin是贻贝免疫系统的重要组成部分,对其结构与功能的研究表明,其序列中连接两段β-折叠的发夹区域是其抗菌功能的关键所在。为验证该区域是否具有抗菌活性,通过对厚壳贻贝Mytilus coruscus抗菌肽Mytilin进行空间结构模拟,选取其中β-发夹部分肽段,采用了固相化学合成的方法合成了两条10肽,分别命名为Mytilin Derived Peptide-1(MDP-1)和Mytilin Derived Peptide-2(MDP-2)。高效液相色谱以及质谱检测结果表明,合成是成功的。抗菌谱研究表明,MDP-1和MDP-2对革兰氏阳性菌、阴性菌以及真菌均具有明显的抑制作用,同时,合成的MDP由于序列短且有两对二硫键,因此对于温度及人血浆均表现出很强的稳定性。上述研究结果为深入了解厚壳贻贝抗菌肽Mytilin的抗菌机制以及在此基础上开发具有应用价值的新型抗菌肽奠定了基础。     

Structure‐activity relationship studies of shortened analogues of the antimicrobial peptide EeCentrocin 1 from the sea urchin Echinus esculentus

EeCentrocin 1 is a potent antimicrobial peptide isolated from the marine sea urchin Echinus esculentus. The peptide has a hetero‐dimeric structure with the antimicrobial activity confined in its largest monomer, the heavy chain (HC), encompassing 30 amino acid residues. The aim of the present study was to develop a shorter drug lead peptide using the heavy chain of EeCentrocin 1 as a starting scaffold and to perform a structure‐activity relationship study with sequence modifications to optimize antimicrobial activity. The experiments consisted of 1) truncation of the heavy chain, 2) replacement of amino acids unfavourable for in vitro antimicrobial activity, and 3) an alanine scan experiment on the truncated and modified heavy chain sequence to identify essential residues for antimicrobial activity. The heavy chain of EeCentrocin 1 was truncated to less than half its initial size, retaining most of its original antimicrobial activity. The truncated and optimized lead peptide ( P6 ) consisted of the 12 N‐terminal amino acid residues from the original EeCentrocin 1 HC sequence and was modified by two amino acid replacements and a C‐terminal amidation. Results from the alanine scan indicated that the generated lead peptide ( P6 ) contained the optimal sequence for antibacterial activity, in which none of the alanine scan peptides could surpass its antimicrobial activity. The lead peptide ( P6 ) was also superior in antifungal activity compared to the other peptides prepared and showed minimal inhibitory concentrations (MICs) in the low micromolar range. In addition, the lead peptide ( P6 ) displayed minor haemolytic and no cytotoxic activity, making it a promising lead for further antimicrobial drug development.     

Antimicrobial activity,bactericidal mechanism and LPS‐neutralizing activity of the cell‐penetrating peptide pVEC and its analogs

pVEC is a cell‐penetrating peptide derived from the murine vascular endothelial‐cadherin protein. To evaluate the potential of pVEC as antimicrobial peptide (AMP), we synthesized pVEC and its analogs with Trp and Arg/Lys substitution, and their antimicrobial and lipopolysaccharide (LPS)‐neutralizing activities were investigated. pVEC and its analogs displayed a potent antimicrobial activity (minimal inhibitory concentration: 4–16 μM) against Gram‐positive and Gram‐negative bacteria but no or less hemolytic activity (less than 10% hemolysis) even at a concentration of 200 μM. These peptides induced a near‐complete membrane depolarization (more than 80%) at 4 μM against Staphylococcus aureus and a significant dye leakage (35–70%) from bacterial membrane‐mimicking liposome at a concentration as low as 1 μM. The fluorescence profiles of pVEC and its analogs in dye leakage from liposome and membrane depolarization were similar to those of a frog‐derived AMP, magainin 2. These results suggest that pVEC and its analogs kill bacteria by forming a pore or ion channel in the cytoplasmic membrane. pVEC and its analogs significantly inhibited nitric oxide production or tumor necrosis factor‐α release in LPS‐stimulated mouse macrophage RAW264.7 cells at 10 to 50 μM, in which RAW264.7 were not damaged. Taken together, our results suggest that pVEC and its analogs with potent antimicrobial and LPS‐neutralizing activities can serve as AMPs for the treatment of microbial infection and sepsis. Copyright © 2011 European Peptide Society and John Wiley & Sons, Ltd.     

Synthesis,spectral properties and antimicrobial activity of a new cationic water‐soluble pH‐dependent poly(propylene imine) dendrimer modified with 1,8‐naphthalimides

A three‐step synthesis was implemented to prepare a quaternary ammonium functionalized blue fluorescent poly(propylene imine) dendrimer modified with pyridinium salt of 4‐acylamino‐1,8‐naphthalimide. The new cationic dendrimer absorbs in the ultraviolet light region and emits blue fluorescence. Its spectral characteristics in organic solvents and in an aqueous solution were studied. The influence of pH on the fluorescence intensity of the dendrimer was established with regard to its use as a pH sensor. The effect of hydroxyl ions on the absorption and fluorescence spectra in dry N,N‐dimethylformamide was also investigated. The antimicrobial activity of the dendrimer was assessed against model pathogenic microorganisms in agar, liquid medium, and after its deposition on cotton fabric.     

Impact of pectin esterification on the antimicrobial activity of nisin‐loaded pectin particles

The relationship between pectin structure and the antimicrobial activity of nisin‐loaded pectin particles was examined. The antimicrobial activity of five different nisin‐loaded pectin particles, i.e., nisin‐loaded high methoxyl pectin, low methoxyl pectin, pectic acid, dodecyl pectin with 5.4 and 25% degree of substitution were tested in the pH range of 4.0–7.0 by agar‐diffusion assay and agar plate count methods. It was found that the degree of esterification of carboxyl group of galacturonic acid in pectin molecule is important for the antimicrobial activity of nisin‐loaded pectin particles. Nisin‐loaded particles prepared using pectic acid or the pectin with low degree of esterification exhibit higher antimicrobial activity than nisin‐loaded high methoxyl pectin particles. Pectins with free carboxyl groups or of low degree of esterification are the most suitable for particles preparation. Moreover, nisin‐loaded pectin particles were active at close to neutral or neutral pH values. Therefore, they could be effectively applied for food preservation. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 33:245–251, 2017     

Considerations for the process development of insect‐derived antimicrobial peptide production

Antimicrobial peptides (AMPs) could evolve into new therapeutic lead molecules against multi‐resistant bacteria. As insects are a rich source of AMP, the identification and characterization of insect‐derived AMPs is particularly emphasized. One challenge of bringing these molecules into market, e.g., as a drug, is to develop a cost‐efficient large‐scale production process. Due to the fact that a direct AMP isolation from insects is not economical and that chemical synthesis is recommended for peptide sizes below 40 amino acids, a viable option is heterologous AMP production. Therefore, previous knowledge concerning the expression of larger proteins can be adapted, but due to the AMP nature (e.g., small size, bactericide) additional challenges have to be faced during up and downstream processing. Nonetheless the bottleneck for large‐scale AMP production is the same as for proteins; mainly the downstream process. This review introduces opportunities for insect‐derived AMP production, like the choice of the expression system (based on previously derived data), depending on the AMP nature, as well as new purification strategies like elastin‐like peptide/intein based purification strategies. All of these aspects are discussed with regard to large‐scale processes and costs. © 2014 American Institute of Chemical Engineers Biotechnol. Prog., 31:1–11, 2015