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Peptides and proteins are important bioorganic compounds in nature, among which a special place is occupied by antimicrobial substances. There are more than 2000 different antimicrobial peptides (AMPs) produced by a variety of living organisms. Bacteriocins produced by bacteria are the minor group, whose chemical structures are most complicated among all AMPs. The review summarized the main points related to antimicrobial action of the bacteriocins including steps of peptide's interaction with bacterial membranes and details of membrane damaging. The membrane-disordered bacteriocins were described in accordance with structural-functional relationships.

The introduction of azole heterocycles into a peptide backbone is the principal step in the biosynthesis of numerous compounds with therapeutic potential. One of them is microcin B17, a bacterial topoisomerase inhibitor whose activity depends on the conversion of selected serine and cysteine residues of the precursor peptide to oxazoles and thiazoles by the McbBCD synthetase complex. Crystal structures of McbBCD reveal an octameric B 4 C 2 D 2 complex with two bound substrate peptides. Each McbB dimer clamps the N-terminal recognition sequence, while the C-terminal heterocycle of the modified peptide is trapped in the active site of McbC. The McbD and McbC active sites are distant from each other, which necessitates alternate shuttling of the peptide substrate between them, while remaining tethered to the McbB dimer. An atomic-level view of the azole synthetase is a starting point for deeper understanding and control of biosynthesis of a large group of ribosomally synthesized natural products., (Copyright © 2018 The Author(s). Published by Elsevier Inc. All rights reserved.)

The structure of the individual peptides of the two-peptide bacteriocin plantaricin S, an antimicrobial peptide produced by a Lactobacillus plantarum strain, has been determined in DPC micelles. The two peptides of plantaricin S, Pls-α and Pls-β, form an α-helix from and including residue 8 to 24 with a less structured region around residue 16-19 and an amphiphilic α-helix from and including residue 7 to 23, respectively. Activity assays on single amino acid-substituted GxxxG and GxxxG-like motifs show that substituting the Ser and Gly residues in the G 9 xxxG 13 motif in Pls-α and the S 17 xxxG 21 motif in Pls-β reduced or drastically reduced the antimicrobial activity. The two-peptide bacteriocin muricidin contains GxxxG-like motifs at similar positions and displays 40-50% amino acid identity with plantaricin S. Activity assays of combinations of the peptides that constitute the bacteriocins plantaricin S and muricidin show that some combinations are highly active. Furthermore, sequence alignments show that the motifs important for plantaricin S activity align with identical motifs in muricidin. Based on sequence comparison and activity assays, a membrane-inserted model of plantaricin S in which the two peptides are oriented antiparallel relative to each other and where the GxxxG and GxxxG-like motifs important for activity come close in space, is proposed.

The antimicrobial peptide bactofencin A is an unmodified non-pediocin-like bacteriocin that inhibits several clinically relevant pathogens, including Listeria monocytogenes and Staphylococcus aureus. Here we report the synthesis and structure-activity relationship studies of bactofencin A and novel analogues thereof. Synthetic bactofencin A was a potent inhibitor of L. monocytogenes (MIC = 8.0 μM) and S. aureus (MIC = 4.0 μM), similar to the bacteriocin produced naturally by Lactobacillus salivarius. Of particular interest is the fact that linear analogues lacking the disulfide bond found in bactofencin A were as potent and also active against several strains of methicillin-resistant S. aureus (MRSA) and one strain of vancomycin-resistant S. aureus (VRSA). Supported by the structure-activity relationship study, investigation of the interaction of bactofencin A with bacterial membrane by molecular dynamics simulations showed the importance of the positively charged N-terminal tail for peptide-membrane interaction. These results suggest that the C-terminal macrocycle is involved in target protein binding and bacterial growth inhibition.

Secondary metabolites are a range of bioactive compounds yielded by bacteria, fungi and plants, etc. The published archaea genomic data provide the opportunity for efficient identification of secondary metabolite biosynthetic gene clusters (BGCs) by genome mining. However, the study of secondary metabolites in archaea is still rare. By using the antiSMASH, we found two main putative secondary metabolite BGCs, bacteriocin and terpene in 203 Archaea genomes. Compared with the genomes of Euryarchaeota that usually lives in less complexity of environment, the genomes of Crenarchaeota usually contained more abundant bacteriocin. In these archaea genomes, we also found the positive correlation between the abundance of bacteriocin and the abundance of CRISPR spacer, suggesting the bacteriocin might be a crucial component of the innate immune system that defense the microbe living in the common environment. The structure analysis of the bacteriocin gene clusters gave a clue that the assisted genes located at the edge of clusters evolved faster than the core biosynthetic genes. To the best of our knowledge, we are the first to systematically explore the distribution of secondary metabolites in archaea, and the investigation of the relationship between BGC and CRISPR spacer expands our understanding of the evolutionary dynamic of these functional molecules., (Copyright © 2018. Published by Elsevier Ltd.)

Aim: Bacteriocins are considered as promising alternatives to antibiotics against infections. In this study, the plantaricins (Pln) A, E, F, J and K were investigated for their antimicrobial activity against Staphylococcus epidermidis., Materials & Methods: The effects on membrane integrity were studied using liposomes and viable bacteria, respectively., Results: We show that PlnEF and PlnJK caused rapid and significant lysis of S. epidermidis, and induced lysis of liposomes. The PlnEF and PlnJK displayed similar mechanisms by targeting and disrupting the bacterial cell membrane. Interestingly, Pln enhanced the effects of different antibiotics by 30- to 500-fold., Conclusion: This study shows that Pln in combination with low concentrations of antibiotics is efficient against S. epidermidis and may be developed as potential treatment of infections.

We report crystal structures of the antibacterial lasso peptides microcin J25 (MccJ25) and capistruin (Cap) bound to their natural enzymatic target, the bacterial RNA polymerase (RNAP). Both peptides bind within the RNAP secondary channel, through which NTP substrates enter the RNAP active site, and sterically block trigger-loop folding, which is essential for efficient catalysis by the RNAP. MccJ25 binds deep within the secondary channel in a manner expected to interfere with NTP substrate binding, explaining the partial competitive mechanism of inhibition with respect to NTPs found previously [Mukhopadhyay J, Sineva E, Knight J, Levy RM, Ebright RH (2004) Mol Cell 14:739-751]. The Cap binding determinant on RNAP overlaps, but is not identical to, that of MccJ25. Cap binds further from the RNAP active site and does not sterically interfere with NTP binding, and we show that Cap inhibition is partially noncompetitive with respect to NTPs. This work lays the groundwork for structure determination of other lasso peptides that target the bacterial RNAP and provides a structural foundation to guide lasso peptide antimicrobial engineering approaches., Competing Interests: The authors declare no conflict of interest., (Copyright © 2019 the Author(s). Published by PNAS.)

Microorganisms are able to produce hundreds of unique chemical structures that can be effectively used by the human beings on their own benefit using the products in the chemical industry. Bacteria belonging to Bacillus genera are very good chemical factories capable to synthesize different compounds with a wide variety of activities. In this review, we try to review the compounds with their respective biological activities produced by different species of Bacillus., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

Lantibiotics, an important group of ribosomally synthesized peptides, represent an important arsenal of novel promising antimicrobials showing high potency in fighting against the prevalence of antibiotic resistance among microbial pathogens. However, due to the lack of high throughput strategies for the isolation and identification of these compounds, our information regarding their structure and especially sequence-based properties is far from complete. Therefore, in the present study, a comprehensive sequence-based analysis of these peptides was performed with the help of machine learning approach together with a feature selection technique. Meanwhile, an attempt to develop an accurate computational model for prediction of lantibiotics was made via constructing two datasets of 280 and 190 lantibiotic and non-lantibiotic antimicrobial peptide sequences, respectively. Based on the conducted approach and as a result of our search for a subset of relevant features of lantibiotics, particular types of sequenced-based features were observed to be preferred in lantibiotics, the knowledge-based implementation of which can be used as strategies for lantibiotic bioengineering purposes. Moreover, a SMO-based classifier was developed for the prediction of lantibiotics with the accuracy and specificity values of 88.5% and 94%, respectively which shows the great potential of the developed algorithm for the prediction of lantibiotcs. Conclusively, the accurate predictor algorithm as well as the identified sequence-based distinctiveness properties of lantibiotics can give valuable information in both the fields of lantibiotic discovery and bioengineering., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Antimicrobial peptides (AMPs) from prokaryotic source also known as bacteriocins are ribosomally synthesized by bacteria belonging to different eubacterial taxonomic branches. Most of these AMPs are low molecular weight cationic membrane active peptides that disrupt membrane by forming pores in target cell membranes resulting in cell death. While these peptides known to exhibit broad-spectrum antimicrobial activity, including antibacterial and antifungal, they displayed minimal cytotoxicity to the host cells. Their antimicrobial efficacy has been demonstrated in vivo using diverse animal infection models. Therefore, we have discussed some of the promising peptides for their ability towards potential therapeutic applications. Further, some of these bacteriocins have also been reported to exhibit significant biological activity against various types of cancer cells in different experimental studies. In fact, differential cytotoxicity towards cancer cells as compared to normal cells by certain bacteriocins directs for a much focused research to utilize these compounds as novel therapeutic agents. In this review, bacteriocins that demonstrated antitumor activity against diverse cancer cell lines have been discussed emphasizing their biochemical features, selectivity against extra targets and molecular mechanisms of action.

Background: The emergence of serious issues of multidrug resistance in the past few years have enforced the use of bacteriocins for combating infections. Threat posed to public health by various multidrug resistant (MDR) organisms can be resolved by discovering new antimicrobial proteins with broad spectrum of inhibition., Results: In the current study, Bacteriocin (BAC-IB17) produced by Bacillus subtilis KIBGE-IB17 is found to be effective against different strains of methicillin resistant Staphylococcus aureus (MRSA). The approximate molecular mass of BAC-IB17 is 10.7 kDa. This unique bacteriocin is found to be highly thermostable and pH stable in nature. It also showed its stability against various heavy metals, organic solvents, surfactants and proteolytic enzymes. Amino acid profile of BAC-IB17 clearly showed that this protein mainly consists of non-polar and basic amino acids whereas; some acidic amino acids were also detected. Sequence of first 15 amino acid residues obtained from N-terminal sequencing of BAC-IB17 were NKPEALVDYTGVXNS., Conclusions: The anti-MRSA property of purified bacteriocin may be used to prevent the spread of MRSA infections. Remarkable features of BAC-IB17 suggests its applications in various pharmaceutical and food industries as it can function under a variety of harsh environmental conditions.

In previous work we have isolated and identified a new strain called Enterococcus faecium FL31. The active compound secreted by this strain, "BacFL31", has been purified and characterized. In the present study, safety aspect, assessed by microbiological and molecular tests, demonstrated that Enterococcus faecium FL31 was susceptible to relevant antibiotics, free of hemolytic, gelatinase, DNase, and lipase activities. In addition, it did not harbor virulence and antibiotic resistance genes. Combined SYTOX Green dye and UV-absorbing experiments, along with released extracellular potassium and transmembrane electrical potential measurements, showed that pure BacFL31 at a concentration of 1×MIC (50  μ g/mL) could damage cytoplasmic membrane of the pathogen Listeria monocytogenes ATCC19117. The same concentration causes the leakage of its intracellular constituents and leads to the destruction of this pathogenic microorganism. In summary, this work reflected characteristics of Enterococcus faecium FL31 strain and its bacteriocin in terms of functional and safety perspectives.

Bacteria use various strategies to compete in an ecological niche, including the production of bacteriocins. Bacteriocins are ribosomally synthesized antibacterial peptides, and it has been postulated that the majority of Gram-positive bacteria produce one or more of these natural products. Bacteriocins can be used in food preservation and are also considered as potential alternatives to antibiotics. The majority of bacteriocins from Gram-positive bacteria had been traditionally divided into two major classes, namely lantibiotics, which are post-translationally modified bacteriocins, and unmodified bacteriocins. The last decade has seen an expanding number of ribosomally synthesized and post-translationally modified peptides (RiPPs) in Gram-positive bacteria that have antibacterial activity. These include linear azol(in)e-containing peptides, thiopeptides, bottromycins, glycocins, lasso peptides and lipolanthines. In addition, the three-dimensional (3D) structures of a number of modified and unmodified bacteriocins have been elucidated in recent years. This review gives an overview on the structural variety of bacteriocins from Gram-positive bacteria. It will focus on the chemical and 3D structures of these peptides, and their interactions with receptors and membranes, structure-function relationships and possible modes of action.

Non-invasive imaging of apoptosis in tumors induced by chemotherapy is of great value in the evaluation of therapeutic efficiency. In this study, we report the synthesis, characterization, and utilization of radionuclide technetium-99m ( 99m Tc)-labeled dendrimer-entrapped gold nanoparticles (Au DENPs) for targeted SPECT/CT imaging of chemotherapy-induced tumor apoptosis. Generation five poly(amidoamine) (PAMAM) dendrimers (G5.NH 2 ) were sequentially conjugated with 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA), polyethylene glycol (PEG) modified duramycin, PEG monomethyl ether, and fluorescein isothiocyanate (FI) to form the multifunctional dendrimers, which were then utilized as templates to entrap gold nanoparticles. Followed by acetylation of the remaining dendrimer surface amines and radiolabeling of 99m Tc, the SPECT/CT dual mode nanoprobe of tumor apoptosis was constructed. The developed multifunctional Au DENPs before and after 99m Tc radiolabeling were well characterized. The results demonstrate that the multifunctional Au DENPs display favorable colloidal stability under different conditions, own good cytocompatibility in the given concentration range, and can be effectively labeled by 99m Tc with high radiochemical stability. Furthermore, the multifunctional nanoprobe enables the targeted SPECT/CT imaging of apoptotic cancer cells in vitro and tumor apoptosis after doxorubicin (DOX) treatment in the established subcutaneous tumor model in vivo. The designed duramycin-functionalized Au DENPs might have the potential to be employed as a nanoplatform for the detection of apoptosis and early tumor response to chemotherapy.

Mannose phosphotransferase system (Man-PTS) serves as a receptor for several bacteriocins in sensitive bacterial cells, namely subclass IIa bacteriocins (pediocin-like; pediocins) and subclass IId ones - lactococcin A (LcnA), lactococcin B (LcnB) and garvicin Q (GarQ). Here, to identify the receptor for three other narrow-spectrum subclass IId bacteriocins - garvicins A, B and C (GarA-C) Lactococcus garvieae mutants resistant to bacteriocins were generated and sequenced to look for mutations responsible for resistance. Spontaneous mutants had their whole genome sequenced while in mutants obtained by integration of pGhost9::ISS1 regions flanking the integration site were sequenced. For both types of mutants mutations were found in genes encoding Man-PTS components IIC and IID indicating that Man-PTS likely serves as the receptor for these bacteriocins as well. This was subsequently confirmed by deletion of the man-PTS operon in the bacteriocin-sensitive L. garvieae IBB3403, which resulted in resistant cells, and by heterologous expression of appropriate man-PTS genes in the resistant Lactococcus lactis strains, which resulted in sensitive cells. GarA, GarB, GarC and other Man-PTS-targeting bacteriocins differ in the amino acid sequence and activity spectrum, suggesting that they interact with the receptor through distinct binding patterns. Comparative analyses and genetic studies identified a previously unrecognized extracellular loop of Man-PTS subunit IID (γ+) implicated in the L. garvieae sensitivity to the bacteriocins studied here. Additionally, individual amino acids localized mostly in the sugar channel-forming transmembrane parts of subunit IIC or in the extracellular parts of IID likely involved in the interaction with each bacteriocin were specified. Finally, template-based 3D models of Man-PTS subunits IIC and IID were built to allow a deeper insight into the Man-PTS structure and functioning.

Glycocins (glycosylated bacteriocins) are a family of ribosomally synthesized and post-translationally modified peptides with antimicrobial activities against pathogens of interest, including methicillin-resistant Staphylococcus aureus, representing a promising source of new antibiotics. Glycocins are still largely underexplored, and thus far, only six glycocins are known. Here, we used genome mining to identify 50 putative glycocin biosynthetic gene clusters and then chose six of them with distinct features for further investigation. Through two rounds of plug-and-play pathway refactoring and expression in Escherichia coli BL21(DE3), four systems produced novel glycocins. Further structural characterization revealed that one of them, which belongs to the enterocin 96-type glycocins, was diglucosylated on a single serine. The other three compounds belong to the SunA/ThuA-type glycocins and exhibit a antimicrobial spectrum narrower than that of sublancin, the best characterized member in this group, even though they share a similar disulfide topology and glycosylation. Further evaluation of their bioactivities with free glucose at high concentrations suggested that their antimicrobial mechanisms might be both glycocin- and species-specific. These glycocins with distinct features significantly broaden our knowledge and may lead to the discovery of new classes of antibiotics.

A novel bacteriocin-like protein and its structural gene (rap) were identified from Rhodococcus erythropolis JCM 2895. The rapA and B genes are located on a 5.4-kb circular plasmid, and were obtained using a modified suppression-subtractive hybridization method. The rapA and B genes were heterologously expressed in Rhodococcus sp. or Escherichia coli, and then characterized. The results indicated that RapA is a small, water-soluble, heat-stable antimicrobial protein, and that RapB is an immunity protein against RapA, estimated to be located on the cell membrane. RapA showed antimicrobial activity particularly against R. erythropolis, and the activity persisted even after SDS-PAGE analysis. For the heterologous expressed RapA protein, N-terminal amino acid sequence was also confirmed. This is the first report of a bacteriocin-like substance obtained from the genus Rhodococcus.

Bacteriocins are ribosomally synthesized antimicrobial proteins/peptides. They are of great interest in the food processing industries as potential natural preservative agent to control food-borne pathogens. Bacillus spp. are one among the potential probiotics receiving more attention since they produce a broad spectrum of antimicrobial bioactive peptides. In this study, a small-scale medium composition and bioprocessing parameters were statistically optimized to increase the yield of bacteriocin namely cerein from Bacillus cereus NS02 showing antagonism against a wide range of food-borne pathogens. The cerein was partially purified, characterized, and evaluated for their optimal reaction condition. It was subjected to surface adsorption onto food-grade silica to evaluate its maximal adsorption, reached at 4 h, 40 °C, pH 6-7, and at the initial concentration of 200 AU mL -1 . The effectiveness of silica-adsorbed and silica-free cerein was checked in Listeria monocytogenes inoculated fresh apple juice and demonstrated biopreservative activity. In juice treated with silica-cerein, the colony forming unit (CFU) was found to be less in count on the 15th day of storage at 4 °C whereas, free-cerein was found to contain 3.8 log CFU mL -1 . While, on the same day of storage, the control juice contained the strength of 14.6 log CFU mL -1 . Based on the above, this study concludes that the identified heat stable low molecular weight peptide cerein from B. cereus NS02 could serve as a potential biopreservative with effective antilisterial activity in the food system. However, a more detailed study is required to determine if their quality change especially the effect of cerein in organoleptic and nutritional properties of food beyond their addition is necessary, before it is to be exploited as an ecofriendly biopreservative.

Aims: The aim of this study was to develop a plantaricin BM-1, a typical IIa bacteriocin produced by Lactocacillus plantarumBM-1, for active polyvinylidene chloride (PVDC) films and to determine the antimicrobial effect of plantaricin BM-1 incorporated into a PVDC film on fresh pork during 7 days of storage at 4°C., Methods and Results: Plantaricin BM-1 solutions (20 480 AU ml -1 ) that absorbed into the PVDC film increased gradually and reached maximum volumes during exposure for up to 20 h. When soaked in water, the released amount of plantaricin BM-1 from the active PVDC film reached a maximum at 20 h. The plantaricin BM-1 active PVDC film had an obvious antilisterial effect in culture medium and fresh pork inoculated with Listeria monocytogenes. Furthermore, plantaricin BM-1-incorporated PVDC film was also significantly (P < 0·01) reduced to aerobic counts of approximately 1·5 log 10 CFU per g after 7 days of storage at 4°C in pork meat, and the pH and total volatile basic nitrogen of pork meat were significantly (P < 0·01, P < 0·05) lower than those of the control., Conclusion: Plantaricin BM-1 active film has an excellent effect to prolong the shelf life of pork meat during cold storage., Significance and Impact of the Study: The results of this study suggest a potential application of bacteriocin active film on meat preservation., (© 2018 The Society for Applied Microbiology.)

Bacteriocins are bioactive antimicrobial peptides synthesized in the ribosome of numerous bacteria and released extracellularly. Bacteriocins have the ability to kill or inhibit the growth of prokaryotes and could potentially be useful against pathogens and antibiotic-resistant strains of bacteria. The antimicrobial mechanisms and relatively narrow killing spectrums of bacteriocins distinguish them from traditional broad-spectrum antibiotics, making them possible candidates to replace antibiotics in the future. In recent years, researchers have discovered many new bacteriocins from different sources. However, the discovery of new bacteriocins involves complicated screening, identification, purification, and characterization processes. This review describes the strategies currently employed for screening, identification, purification, and characterization of bacteriocins. Approximately 40 novel bacteriocins are reviewed in this paper. Our conclusions about the research process will guide the development of novel methods and allow for the faster, easier, and more efficient discovery of novel bacteriocins., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Modern large-scale agricultural practices that incorporate high density farming with subtherapeutic antibiotic dosing are considered a major contributor to the rise of antibiotic-resistant bacterial infections of humans with species of Salmonella being a leading agriculture-based bacterial infection. Microcin J25, a potent and highly stable antimicrobial peptide active against Enterobacteriaceae, is a candidate antimicrobial against multiple Salmonella species. Emerging evidence supports the hypothesis that the composition of the microbiota of the gastrointestinal tract prevents a variety of diseases by preventing infectious agents from proliferating. Reducing clearance of off-target bacteria may decrease susceptibility to secondary infection. Of the Enterobacteriaceae susceptible to microcin J25, Escherichia coli are the most abundant within the human gut. To explore the modulation of specificity, a collection of 207 mutants encompassing 12 positions in both the ring and loop of microcin J25 was built and tested for activity against Salmonella and E. coli strains. As has been found previously, mutational tolerance of ring residues was lower than loop residues, with 22% and 51% of mutations, respectively, retaining activity toward at least one target within the target organism test panel. The multitarget screening elucidated increased mutational tolerance at position G2, G3, and G14 than previously identified in panels composed of single targets. Multiple mutations conferred differential response between the different targets. Examination of specificity differences between mutants found that 30% showed significant improvements to specificity toward any of the targets. Generation and testing of a combinatorial library designed from the point-mutant study revealed that microcin J25 I13T reduces off-target activity toward commensal human-derived E. coli isolates by 81% relative to Salmonella enterica serovar Enteritidis. These in vitro specificity improvements are likely to improve in vivo treatment efficacy by reducing clearance of commensal bacteria in the gastrointestinal tract of hosts., (© 2018 Wiley Periodicals, Inc.)

Duramycin is a heavily post-translationally modified peptide that binds phosphatidylethanolamine. It has been investigated as an antibiotic, an inhibitor of viral entry, a therapeutic for cystic fibrosis, and a tumor and vasculature imaging agent. Duramycin contains a β-hydroxylated Asp (Hya) and four macrocycles, including an essential lysinoalanine (Lal) cross-link. The mechanism of Lal formation is not known. Here we show that Lal is installed stereospecifically by DurN via addition of Lys19 to a dehydroalanine. The structure of DurN reveals an unusual dimer with a new fold. Surprisingly, in the structure of duramycin bound to DurN, no residues of the enzyme are near the Lal cross-link. Instead, Hya15 of the substrate makes interactions with Lal, suggesting it acts as a base to deprotonate Lys19 during catalysis. Biochemical data suggest that DurN preorganizes the reactive conformation of the substrate, such that the Hya15 of the substrate can serve as the catalytic base for Lal formation.

Discovery of a novel bacteriocin is always an event in sciences, since cultivation of most bacterial species is a general problem in microbiology. This statement is reflected by the fact that number of bacteriocins is smaller for tenfold comparing to known antimicrobial peptides. We cultivated Enterococcus faecium on simplified medium to reduce amount of purification steps. This approach allows to purify the novel heavy weight bacteriocin produced by E. faecium ICIS 7. The novelty of this bacteriocin, named enterocin-7, was confirmed by N-terminal sequencing and by comparing the structural-functional properties with available data. Purified enterocin-7 is characterized by a sequence of amino acid residues having no homology in UniProt/SwissProt/TrEMBL databases: NH2 - Asp - Ala - His - Leu - Ser - Glu - Val - Ala - Glu - Arg - Phe - Glu - Asp - Leu - Gly. Isolated thermostable protein has a molecular mass of 65 kDa, which allows it to be classified into class III in bacteriocin classification schemes. Enterocin-7 displayed a broad spectrum of activity against some Gram-positive and Gram-negative microorganisms. Fluorescent microscopy and spectroscopy showed the permeabilizing mechanism of the action of enterocin-7, which is realized within a few minutes.

Bactofencin A is a novel class IId bacteriocin, produced by the intestinal isolate Lactobacillus salivarius DPC6502, which has potent activity against medically significant pathogens including Staphylococcus aureus. This bacteriocin is unusual in that it has a highly cationic N terminus and a single disulfide bond between Cys7 and Cys22, resulting in a large C terminal loop. In this study, a library of synthetic bactofencin A variants were screened against the mastitis isolate, S. aureus DPC5246, to identify key residues responsible for activity. It was apparent that substituting either cysteine of the disulfide bond with either serine or alanine significantly reduced the activity of the bacteriocin, confirming the importance of the C terminal loop. Substituting N terminal amino acids with alanine had no effect on activity, whereas sequential removal of the N terminal positively charged residues resulted in an increasingly inactive peptide. A complete (synthetic) alanine scanning analysis revealed that the residues between Val9 and Gly17 were most affected by substitution suggesting that this area has a major influence on the potency of the bacteriocin. Substituting residues in the loop region between Cys7 and Cys22 for D-amino acid equivalents had a more detrimental effect on activity than L-alanine substitutions. Specifically Y10A, N11A, P15A and T16A are active at 4, 16, 1 and 16 μM respectively while their D equivalents were inactive at 1000 μM, the highest concentration tested. Ultimately, this study identifies the critical features in the primary structure of the bacteriocin which gives it such potent activity against pathogenic staphylococci.

Metallic nanoparticles have a substantial scientific interest because of their distinctive physicochemical and antimicrobial properties and the emergence of multidrug resistant pathogens could unlock the potential of nanoparticles to combat infectious diseases. The aim of the current study is to enhance the antibacterial potential of purified bacteriocin by combining bacteriocin and antibacterial silver nanoparticles (AgNPs). Hence, the interaction of natural antimicrobial compounds and antibacterial nanoparticles can be used as a potential tool for combating infectious diseases. In this study, a green, simple and effective approach is used to synthesize antibacterial AgNPs using fungal exopolysaccharide as both a reducing and stabilizing agent. The AgNPs were characterized by spectroscopic analysis, Scanning Electron Microscopy (SEM), Energy Dispersive X-ray spectroscopy (EDX) and Dynamic Light Scattering (DLS). Furthermore, the synergistic effect of bacteriocin-AgNPs was determined against pathogenic strains. The histogram of AgNPs indicated well-dispersed, stabilized and negatively charged particles with variable size distribution. The combination of bacteriocin with nanoparticles found to be more effective due to broad antibacterial potential with possibly lower doses. The current study is imperative to provide an alternative for the chemical synthesis of silver nanoparticles. It showed environmental friendly and cost effective green synthesis of antibacterial nanoparticles., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Mutacin 1140 belongs to the epidermin family of type AI lantibiotics. This family has a broad spectrum of activity against Gram-positive bacteria. The binding of mutacin 1140 to lipid II leads to the inhibition of cell wall synthesis. Pharmacokinetic experiments with type AI lantibiotics are generally discouraging for clinical applications due to the short half-life of these compounds. The unprotected dehydrated and protease-susceptible residues outside the lanthionine rings may play a role in the short half-life in physiological settings. Previous mutagenesis work on mutacin 1140 has been limited to the lanthionine-forming residues, the C-terminally decarboxylated residue, and single amino acid substitutions at residues Phe1, Trp4, Dha5, and Arg13. To study the importance of the dehydrated (Dha5 and Dhb14) and protease-susceptible (Lys2 and Arg13) residues within mutacin 1140 for stability and bioactivity, each of these residues was evaluated for its impact on production and inhibitory activity. More than 15 analogs were purified, enabling direct comparison of the activities against a select panel of Gram-positive bacteria. The efficiency of the posttranslational modification (PTM) machinery of mutacin 1140 is highly restricted on its substrate. Analogs in the various intermediate stages of PTMs were observed as minor products following single point mutations at the 2nd, 5th, 13th, and 14th positions. The combination of alanine substitutions at the Dha5 and Dhb14 positions abolished mutacin 1140 production, while the production was restored by substitution of a Gly residue at one of these positions. Analogs with improved activity, productivity, and proteolytic stability were identified. IMPORTANCE Our findings show that the efficiency of mutacin 1140 PTMs is highly dependent on the core peptide sequence. Analogs in various intermediate stages of PTMs can be transported by the bacterium, which indicates that PTMs and transport are finely tuned for the native mutacin 1140 core peptide. Only certain combinations of amino acid substitutions at the Dha5 and Dhb14 dehydrated residue positions were tolerated. Observation of glutamylated core peptide analogs shows that dehydrations occur in a glutamate-dependent manner. Interestingly, mutations at positions outside rings A and B, the lipid II binding domain, would interfere with lipid II binding. Purified mutacin 1140 analogs have various activities and selectivities against different genera of bacteria, supporting the effort to generate analogs with higher specificity against pathogenic bacteria. The discovery of analogs with improved inhibitory activity against pathogenic bacteria, increased stability in the presence of protease, and higher product yields may promote the clinical development of this unique antimicrobial compound., (Copyright © 2018 American Society for Microbiology.)

A previously reported gene cluster encoding four nisin-like peptides, three with the same sequence (NsoA1-3) and the unique NsoA4, produced antimicrobial activity in the presence of trypsin after heterologous expression in Lactococcus lactis. Protein extracts were separated by SDS gel electrophoresis or immunoprecipitation using an antibody to the NsoA2 leader. Tryptic peptides observed by LC-MS/MS covered the complete sequence of preNsoA1-3 and part of the leader sequence of preNsoA4 and confirmed the expression and the predicted sequences of the preNsoA peptides. Further, the data revealed that the preNsoA1-3 peptides were partly modified with dehydrations and formation of lanthionine rings. A certain amount of fully modified preNsoA1-3 was observed. Details of modifications of the core peptide and the C-terminal tryptic peptide TATCGCHITGK covering rings D and E indicated that 22% of these preNsoA1-3 peptides were completely modified. A lower amount of ring formation is estimated for rings A-C. Intact masses of immunoprecipitation-derived peptides determined by LC-MS accurately matched the expected preNsoA precursor peptides. The most abundant peptides detected were preNsoA2-3-8H 2 O followed by preNsoA1-8H 2 O and other states of dehydration. The results confirm incomplete processing of preNsoA peptides in the heterologous system, with the formation of a certain amount of fully modified peptides.

The development of alternatives for the use of chemical pesticides for plant disease control is the present-day and ongoing challenge for achieving sustainable agriculture. Pseudomonas fluorescens SF4c, native strain from wheat, produces tailocins (phage-tail-like bacteriocins) with antimicrobial activity against several phytopathogenic strains. We thus investigated the efficacy of foliar application of these bacteriocins to control the bacterial-spot disease in tomato caused by Xanthomonas vesicatoria Xcv Bv5-4a. The disease severity and incidence index were reduced by 44 and 36%, respectively; while the number of viable cells of X. vesicatoria Xcv Bv5-4a decreased after bacteriocin treatment. Furthermore, bacteriocin was effective in reducing bacterial-spot-disease symptoms on tomato fruits even when applied 12 h after infection. Tailocin activity was not affected by abiotic influences such as adjuvant, light and temperature and, biotic factors such as apoplastic-fluids. In contrast, no antibacterial activity of these tailocins was observed when the bacteriocin was exposed to extremely dry conditions. Finally, that no cytotoxic effects on mammalian cells were observed with this representative tailocins is highly significant and demonstrates the safety of such compounds in humans. All these findings indicate that the SF4c tailocins represent an attractive alternative to copper-containing bactericides for use in the control of bacterial spot., (Copyright © 2018 Elsevier GmbH. All rights reserved.)

The aim of this study was to isolate and investigate the bacteriocinogenic and probiotic potential of new Gram-negative isolates. Of 22 bacterial isolates from pig intestine and chicken crops, ten isolates had demonstrated a good activity, and the most potent five strains were identified as four E. coli and one as Proteus sp. No virulence factors were detected for E. coli strains isolated from pig intestine. The semi-purified microcins proved to be resistant to temperature and pH variation, but sensitive to proteolytic enzymes. Of particular interest, strain E. coli P2C was the most potent, free of virulence genes and sensitive to tested antibiotics. Purification procedure revealed the presence of a single pure peak having a molecular mass of 8733.94 Da and matching microcin V (MccV). The sequence obtained by LC-MS/MS confirmed the presence of MccV. Purified MccV showed a good activity against pathogenic coliforms, especially E. coli O 1 K 1 H 7 involved in avian colibacillosis. The present study provides evidence that E. coli strains isolated from pig intestine produce microcin-like substances. E. coli P2C is a safe MccV producer that could be a good candidate for its application as novel probiotic strain to protect livestock and enhance growth performance.

Antimicrobial substances are widely used in many anthropic activities, including sanitary and military services for the human population. These compounds are also known to be used in food production, agricultural activities, and partially correlated industrial sectors. However, there are concerns regarding the link between the abuse of antimicrobial agents in these ambits and the possible detection of antibiotic-resistant microorganisms. Modern food and beverage products are generally found on the market as prepackaged units, with several exceptions. Consequently, positive and negative features of a specific food or beverage should be considered as the result of the synergic action of different components, including the container (or the assembled sum of packaging materials). At present, the meaning of food container also includes the creation and development of new packaging materials that are potentially able to interact with the contained food. "Active" packaging systems can be realized with antimicrobial substances. On the other hand, a careful evaluation of risks and advantages correlated with antimicrobial agents is needed because of possible negative and/or unexpected failures.

Cyanobacteria produce many biologically active metabolites synthesized via nonribosomal synthetic pathways such as cyclic microcystins (MCs) and linear aeruginosins (Aers). The present study aimed to investigate the effects of different MC variants and the newly isolated aerugenosin Aer-865 on macrophages, which represent one of the key effector cells within the innate immune responses. Specifically, our study included RAW 264.7 macrophage activation associated with production of cytotoxic and cytostatic nitric oxide (NO) as well as pro-inflammatory mediators like tumor necrosis factor α (TNFα) and interleukin 6 (IL-6). From the compounds investigated, commonly occurring MC variants (-RR, -YR) and Aer-865 had no significant effects within the non-cytotoxic concentrations tested, i.e. 0.001-1 μM for MCs and 0.1-50 μM for Aer-865. In contrast to known immunoactive MC-LR, the negligible immunomodulatory potential of tested MC congeners could be related to their differences in structure. The knowledge of MC structure-specific activities contributes to the understanding of complex toxicity of different MC variants and most importantly their mixtures. This study is one of the first study that evaluate the effect of larger set of cyanobacterial peptides on macrophages and compare their immunomodulatory potential., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Bacteria under stress produce ribosomally synthesized and post-translationally modified peptides (RiPPs) to target closely related species, such as the lasso peptide microcin J25 (MccJ25). These peptides are also toxic to the producing organisms that utilize dedicated ABC transporters to achieve self-immunity. MccJ25 is exported by the Escherichia coli ABC transporter McjD through a complex mechanism of recognition that has remained elusive. Here, we used biomolecular NMR to study this interaction and identified a region of the toxic peptide that is crucial to its recognition by the ABC transporter. Our study provides evidence that McjD is highly specific to MccJ25 and not to other RiPPs or antibiotics, unlike multidrug ABC transporters. Additionally, we show that MccJ25 is not exported by another natural product ABC transporter. Therefore, we propose that specific interactions between natural product ABC transporters and their substrate provides them with their high degree of specificity. Taken together, these findings suggest that ABC transporters might have acquired structural elements in their binding cavity to recognize and allow promiscuous export of a larger variety of compounds.

The antimicrobial peptide pediocin PA-1 is a class IIa bacteriocin that inhibits several clinically relevant pathogens including Listeria spp. Here we report the synthesis and characterization of whole pediocin PA-1 and novel analogs thereof using a combination of solid- and solution-phase strategies to overcome difficulties due to instability and undesired reactions. Pediocin PA-1 thus synthesized was a potent inhibitor of Listeria monocytogenes (MIC = 6.8 nM), similar to the bacteriocin produced naturally by Pediococcus acidilactici. Of particular interest is that linear analogs lacking both of the disulfide bridges characterizing pediocin PA-1 were as potent. One linear analog was also a strong inhibitor of Clostridium perfringens, another important food-borne pathogen. These results are discussed in light of conformational information derived from circular dichroism, solution NMR spectroscopy and structure-activity relationship studies.

Genome sequencing of Enterococcus faecium M3K31, a strain isolated from griffon vultures, previously revealed the presence of three sequences encoding for bacteriocins, namely enterocin P, enterocin HF, and a SRCAM 602-like bacteriocin. In this work, we describe the SRCAM 602-like bacteriocin that we named faerocin MK. Mature faerocin MK consists of 43 residues and contains an YGNGV-motif and two cysteine residues at positions 10 and 15, consistent with other class IIa bacteriocins. Faerocin MK and SRCAM 602 were chemically synthesized and their scope of activity was tested. Faerocin MK is active against a wide range of Gram-positive organisms while SRCAM 602 was not active against any tested organism. Although both peptides are more structured in trifluoroethanol, faerocin MK has an α-helical character nearly twice that of SRCAM 602. Nucleotide sequence analysis revealed that the faerocin MK precursor is produced with a 28-amino acid signal peptide and that an immunity gene follows the structural faerocin MK gene. Heterologous expression of the faerocin MK operon showed that faerocin MK and its immunity protein were successfully expressed in other organisms. This indicates that the bacteriocin is secreted through the sec pathway.

Leuconostoc pseudomesenteroides 607, isolated from persimmon fruit, was found to have high inhibitory activity against Listeria monocytogenes and several other Gram-positive bacteria. Inhibitory substances were purified from culture supernatant by ion-exchange chromatography, Sep-Pak C 18 cartridge, and reverse-phase high-performance liquid chromatography (RP-HPLC). Two antibacterial peptides were observed during the purification procedures. One of these peptides had a molecular size of 4623.05 Da and a partial N-terminal amino acid sequence of NH 2 -KNYGNGVHxTKKGxS, in which the YGNGV motif is specific for class IIa bacteriocins. A BLAST search revealed that this bacteriocin was similar to leucocin C from Leuconostoc mesenteroides. Leucocin C-specific primers were designed and a single PCR product was amplified. Analysis of the nucleotide sequence has revealed a putative peptide differing by only one amino acid residue from the sequence of leucocin C. No identical peptide or protein has been reported in the literature, and this peptide, termed leucocin C-607, was therefore considered to be a new variant of leucocin C produced by Leuc. pseudomesenteroides 607. Another antibacterial peptide purified from the same culture supernatant had a molecular size of 3007.7 or 3121.97 Da. However, detailed information regarding this second peptide remains to be determined. Distinct characteristics, such as heat stability and inhibitory spectrum, were observed for the two bacteriocins produced by Leuc. pseudomesenteroides 607. These results suggested that Leuc. pseudomesenteroides 607 produces leucocin C-607 along with another unknown bacteriocin.

Lantibiotics are ribosomally synthesized and posttranslationally modified antimicrobial compounds containing lanthionine and methyl-lanthionine residues. Nisin, one of the most extensively studied and used lantibiotics, has been shown to display very potent activity against Gram-positive bacteria, and stable resistance is rarely observed. By binding to lipid II and forming pores in the membrane, nisin can cause the efflux of cellular constituents and inhibit cell wall biosynthesis. However, the activity of nisin against Gram-negative bacteria is much lower than that against Gram-positive bacteria, mainly because lipid II is located at the inner membrane, and the rather impermeable outer membrane in Gram-negative bacteria prevents nisin from reaching lipid II. Thus, if the outer membrane-traversing efficiency of nisin could be increased, the activity against Gram-negative bacteria could, in principle, be enhanced. In this work, several relatively short peptides with activity against Gram-negative bacteria were selected from literature data to be fused as tails to the C terminus of either full or truncated nisin species. Among these, we found that one of three tails (tail 2 [T2; DKYLPRPRPV], T6 [NGVQPKY], and T8 [KIAKVALKAL]) attached to a part of nisin displayed improved activity against Gram-negative microorganisms. Next, we rationally designed and reengineered the most promising fusion peptides. Several mutants whose activity significantly outperformed that of nisin against Gram-negative pathogens were obtained. The activity of the tail 16 mutant 2 (T16m2) construct against several important Gram-negative pathogens (i.e., Escherichia coli , Klebsiella pneumoniae , Acinetobacter baumannii , Pseudomonas aeruginosa , Enterobacter aerogenes ) was increased 4- to 12-fold compared to that of nisin. This study indicates that the rational design of nisin can selectively and significantly improve its outer membrane-permeating capacity as well as its activity against Gram-negative pathogens. IMPORTANCE Lantibiotics are antimicrobial peptides that are highly active against Gram-positive bacteria but that have relatively poor activity against most Gram-negative bacteria. Here, we modified the model lantibiotic nisin by fusing parts of it to antimicrobial peptides with known activity against Gram-negative bacteria. The appropriate selection of peptidic moieties that could be attached to (parts of) nisin could lead to a significant increase in its inhibitory activity against Gram-negative bacteria. Using this strategy, hybrids that outperformed nisin by displaying 4- to 12-fold higher levels of activity against relevant Gram-negative bacterial species were produced. This study shows the power of modified peptide engineering to alter target specificity in a desired direction., (Copyright © 2018 American Society for Microbiology.)

Glycocin F, a bacteriocin produced by Lactobacillus plantarum KW30, is glycosylated with two N-acetyl-d-glucosamine sugars, and has been shown to exhibit a rapid and reversible bacteriostasis on susceptible cells. The roles of certain structural features of glycocin F have not been studied to date. We report here the synthesis of various glycocin F analogues through solid-phase peptide synthesis (SPPS) and native chemical ligation (NCL), allowing us to probe the roles of different structural features of this peptide. Our results indicate that the bacteriostatic activity of glycocin F is controlled by the glycosylated interhelical loop, while the glycosylated flexible tail appears to be involved in localizing the peptide to its cellular target.

Bacillus cereus is an opportunistic pathogen that causes foodborne diseases. We isolated a novel bacteriocin, designated plantaricin GZ1-27, and elucidated its mode of action against B. cereus. Plantaricin GZ1-27 was purified using ammonium sulfate precipitation, gel-filtration chromatography, and RP-HPLC. MALDI-TOF/MS revealed that its molecular mass was 975 Da, and Q-TOF-MS/MS analysis predicted the amino acid sequence as VSGPAGPPGTH. Plantaricin GZ1-27 showed thermostability and pH stability. The antibacterial mechanism was investigated using flow cytometry, confocal laser-scanning microscopy, scanning and transmission electron microscopy, and RT-PCR, which revealed that GZ1-27 increased cell membrane permeability, triggered K + leakage and pore formation, damaged cell membrane integrity, altered cell morphology and intracellular organization, and reduced the expression of genes related to cytotoxin production, peptidoglycan synthesis, and cell division. These results suggest that plantaricin GZ1-27 effectively inhibits B. cereus at both the cellular and the molecular levels and is a potential natural food preservative targeting B. cereus.

Antimicrobial resistance within a wide range of infectious agents is a severe and growing public health threat. Antimicrobial peptides (AMPs) are among the leading alternatives to current antibiotics, exhibiting broad spectrum activity. Their activity is determined by numerous properties such as cationic charge, amphipathicity, size, and amino acid composition. Currently, only around 10% of known AMP sequences have experimentally solved structures. To improve our understanding of the AMP structural universe we have carried out large scale ab initio 3D modeling of structurally uncharacterized AMPs that revealed similarities between predicted folds of the modeled sequences and structures of characterized AMPs. Two of the peptides whose models matched known folds are Lebocin Peptide 1A (LP1A) and Odorranain M, predicted to form β-hairpins but, interestingly, to lack the intramolecular disulfide bonds, cation-π or aromatic interactions that generally stabilize such AMP structures. Other examples include Ponericin Q42, Latarcin 4a, Kassinatuerin 1, Ceratotoxin D, and CPF-B1 peptide, which have α-helical folds, as well as mixed αβ folds of human Histatin 2 peptide and Garvicin A which are, to the best of our knowledge, the first linear αββ fold AMPs lacking intramolecular disulfide bonds. In addition to fold matches to experimentally derived structures, unique folds were also obtained, namely for Microcin M and Ipomicin. These results help in understanding the range of protein scaffolds that naturally bear antimicrobial activity and may facilitate protein design efforts towards better AMPs., (© 2018 The Authors Proteins: Structure, Function, and Bioinformatics Published by Wiley Periodicals, Inc.)

AS-48 is a bacteriocin with potential application as food biopreservative. In order to optimize its use for oral consumption, we assess the impact of gastrointestinal digestion, both in bulk and adsorbed at the air-water interface. Analysis of AS-48 digestion fragments in bulk by SDS-PAGE, RP-HPLC, and MALDI-TOF proves that the previous pepsin exposition promotes digestion by trypsin/chymotrypsin by exposing new cleavage sites. Regarding adsorbed AS-48, the in vitro digestion profile shows that the conformational change undergone by AS-48 upon adsorption affects its digestibility. Gastrointestinal enzymes cleave only susceptible residues, which are oriented into the aqueous phase, while hydrophobic susceptible residues remain undigested. Evaluation of the elasticity of the adsorbed layer confirms also the presence of undigested AS-48. These results are important towards the use of AS-48 in food formulations; assuring that some intact AS-48 resists digestion guarantees its antibacterial activity throughout the gastrointestinal tract., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Lantibiotics are ribosomally synthesized and post-translationally modified peptide natural products that contain thioether cross-links formed by lanthionine and methyllanthionine residues. They exert potent antimicrobial activity against Gram-positive bacteria. We herein report production of analogues of two lantibiotics, lacticin 481 and nisin, that contain nonproteinogenic amino acids using two different strategies involving amber stop codon suppression technology. These methods complement recent alternative approaches to incorporate nonproteinogenic amino acids into lantibiotics.

Bacteriocin LB44 was purified from cell-free supernatant (CFS) of Pediococcus pentosaceus LB44 using activity-guided chromatography techniques. It was stable up to 121 °C, pH 2.0-6.0, sensitive to proteinase K, papain and trypsin, and retained complete activity in the presence of organic solvents tested. The molecular weight of bacteriocin was ∼6 kDa and initial ten amino acid residues (GECGMCXECG) suggested a new compound. The loss in viable cell count and K + ion efflux of target cells of Micrococcus luteus suggested bactericidal activity. The cell membrane of bacteriocin-treated cells was found to be ruptured which was further confirmed by Fourier Transform Infrared (FTIR) analysis suggesting interaction of bacteriocin with phospholipids in cell membrane. It showed broad host-range and inhibited the growth of Lactobacillus delbrueckii NRRL B-4525, L. plantarum NRRL B-4496, L. acidophilus NRRL B-4495, Enterococcus hirae LD3, Weissella confusa LM85, Staphylococcus aureus, Salmonella typhi ATCC 13311, Serratia marcescens ATCC 27137, Pseudomonas aeruginosa ATCC 27853, Proteus vulgaris ATCC 29905, Haloferax larsenii HA1, HA3, HA8, HA9 and HA10. These properties suggested a new bacteriocin from soil isolate P. pentosaceus LB44 which may offers possible applications in food-safety and therapeutics., (Copyright © 2018 Elsevier Inc. All rights reserved.)

The aims of this study were to purify and characterize a bacteriocin produced by a strain of Enterococcus faecalis TG2 and to test the safety of the strain. In this work, the active peptide was purified through precipitation with 70% saturated ammonium sulfate, cation-exchange chromatography, and gel filtration. The specific activity of purified bacteriocin was 30,073.42 AU/mg of protein, which corresponded to a 33.34-fold increase. The molecular mass of the purified bacteriocin was 6.3362 kDa determined by LC-MS/MS. The ten amino acid of N-terminal was MTRSKKLNLR and the ten amino acid of C-terminal was ATGGAAGWKS. The activity of the bacteriocin was unaffected by pH 2-10 and thermostable but was sensitive to proteolytic enzymes. The antimicrobial activity of the bacteriocin was not affected by metal ions. Tween-20, Tween-80, Triton X-100, and EDTA did not affect the bacteriocin activity and SDS was able to increase the activity of bacteriocin. Bacteriocin activity was not lost after treatment by < 8% NaCl. Inhibitory spectrum of the bacteriocin showed a wide range of activities against other lactic acid bacteria, food-spoilage, and food-borne pathogens. Ent. faecalis TG2 was sensitive to tetracycline and erythromycin but resistant to ampicillin, gentamicin, kanamycin, and chloramphenicol. Results from PCR indicated that Ent. faecalis TG2 did not harbor any virulence genes. The study suggests that Ent. faecalis TG2 and its bacteriocin might be used as bio-preservatives in food products.

Purpose: Apoptosis is a key factor in unstable plaques. The aim of this study is to evaluate the utility of visualizing atherosclerotic plaques with radiolabeled duramycin and Annexin V., Procedures: ApoE -/- mice were fed with a high-fat diet to develop atherosclerosis, C57 mice as a control. Using a routine conjugation protocol, highly pure [ 99m Tc]duramycin and [ 99m Tc]Annexin V were obtained, which were applied for in vitro cell assays of apoptosis and in vivo imaging of atherosclerotic plaques in the animal model. Oil Red O staining, TUNEL, hematoxylin-eosin (HE), and CD68 immunostaining were used to evaluate the deposition of lipids and presence of apoptotic macrophages in the lesions where focal intensity positively correlated with the uptake of both tracers., Results: [ 99m Tc]duramycin and [ 99m Tc]Annexin V with a high radiochemical purity (97.13 ± 1.52 and 94.94 ± 0.65 %, respectively) and a well stability at room temperature were used. Apoptotic cells binding activity to [ 99m Tc]duramycin (Kd, 6.92 nM and Bmax, 56.04 mol/10 19 cells) was significantly greater than [ 99m Tc]Annexin V (Kd, 12.63 nM and Bmax, 31.55 mol/10 19 cells). Compared with [ 99m Tc]Annexin V, [ 99m Tc]duramycin bound avidly to atherosclerotic lesions with a higher plaque-to-background ratio (P/B was 8.23 ± 0.91 and 5.45 ± 0.48 at 20 weeks, 15.02 ± 0.23 and 12.14 ± 0.22 at 30 weeks). No plaques were found in C57 control mice. Furthermore, Oil Red O staining showed lipid deposition areas were significantly increased in ApoE -/- mice at 20 and 30 weeks, and TUNEL and CD68 staining confirmed that the focal uptake of both tracers contained abundant apoptotic macrophages., Conclusions: This stable, fast clearing, and highly specific [ 99m Tc]duramycin, therefore, can be useful for the quantification of vulnerable atherosclerotic plaques.

This review attempts to analyze the mechanism of action and immunity of class IIa bacteriocins. These peptides are promising alternative food preservatives and they have a great potential application in medical sciences. Class IIa bacteriocins act on the cytoplasmic membrane of Gram-positive cells dissipating the transmembrane electrical potential by forming pores. However, their toxicity and immunity mechanism remains elusive. Here we discuss the role of the mannose phosphotransferase system (man-PTS) as the receptor for class IIa bacteriocins and the influence of the membrane composition on the activity of these antimicrobial peptides. A model that is consistent with experimental results obtained by different researchers involves the non-specific binding of the bacteriocin to the negatively charged membrane of target bacteria. This step would facilitate a specific binding to the receptor protein, altering its functionality and forming an independent pore in which the bacteriocin is inserted in the membrane. An immunity protein could specifically recognize and block the pore. Bacteriocins function in bacterial ecosystems and energetic costs associated with their production are also discussed. Theoretical models based on solid experimental evidence are vital to understand bacteriocins mechanism of action and to promote new technological developments.

Strains belonging to the genus of Staphylococci, such as Staphylococcus aureus are common pathologic bacteria which may cause nosocomial cross infection and food contamination. Plantaricin EF (PlnEF), a two-peptide nonlantibiotic bacteriocin produced by Lactobacillus plantarum strains, shows great inhibitory effects against Gram-positive Staphylococci strains. To overproduce this two-peptide bacteriocin, plnE and plnF genes were cloned into pET32a (+) vector and heterologously expressed in Escherichia coli by fusion with His6-tag in this study. The purified fusion proteins were cleaved by enterokinase, then PlnE and PlnF peptides without extra amino acids were obtained by a two-step purification method, ultrafiltration centrifuge (10 kDa) followed by a reverse-phase HPLC. Purity of PlnE and PlnF, determined by analytical HPLC, is ∼98%, and their molecular mass confirmed by ESI-MS was 3545.14 Da and 3703.1 Da, respectively. It was found that the two peptides had significant antimicrobial activities against Staphylococcus citreus and Staphylococcus aureus strains and they functioned synergistically. PlnEF exerted its bactericidal activity against Staphylococci strains by permeabilizing the cell membrane, causing influx and efflux of various molecules across the transmembrane barrier, eventually leading to cell death., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Agrocin 108 is a 3'-O-β-D-xylopyranosyl-cytidine-5'-O-phosphodiester of an ascorbate-carbocyclic cyclopentenone analogue, with bacteriocin-like properties. This bacteriocin exhibits orders of magnitude greater than the inhibition zone diameter towards the indicator strain than either ampicillin or streptomycin. It has been isolated from cultures of Rhizobium rhizogenes strain K108. The structure of the agrocin 108 without detail, has been previously published. We now report a detailed structure elucidation, including the hitherto undetermined residual 5'-phospho-diester fragment by a combination of 1D and 2D NMR studies at various pH values in H 2 O/D 2 O, high resolution MS, pKa determination, and chemical degradation.

Nisin is a lantibiotic, a member of a family of polypeptides containing lanthionine with antimicrobial activity. Nisin-producing microorganisms require immunity proteins for self-protection from nisin itself. Lactococcus lactis , a microorganism that synthesizes nisin, has an integral NisFEG ABC transporter and an NisI lipoprotein that function in nisin immunity. Here, we present the crystal structure of the full length of NisI 22-C , a lipid-free form of NisI, determined at 1.9-Å resolution. As with the nuclear magnetic resonance (NMR) structures of the N- and C-terminal domains of NisI, NisI 22-C is composed of N- and C-terminal domains, both of which display a fold similar to that found in SpaI, a lipoprotein with immunity against subtilin in Bacillus subtilis The full-length structure of NisI 22-c reveals a large, deep cleft by the interdomain association, one side of which is occupied by the residues important for immunity. Opposite the cleft, a shallow groove is found where nisin-interacting residues are distributed in the periphery composed of the C-terminal negative patch. Based on a sulfate ion found in the large and deep cleft, a model of NisI in complex with a farnesyl diphosphate backbone of lipid II is proposed, suggesting a mechanism for increasing the chances of encountering nisin., (Copyright © 2018 American Society for Microbiology.)

Lantibiotics are a class of lanthionine-containing, ribosomally synthesized, and posttranslationally modified peptides (RiPPs) produced by Gram-positive bacteria. Salivaricin A2 belongs to the type AII lantibiotics, which are generally considered to kill Gram-positive bacteria by binding to the cell wall precursor lipid II via a conserved ring A structure. Salivaricin A2 was first reported to be isolated from a probiotic strain, Streptococcus salivarius K12, but the structural and bioactivity characterizations of the antibiotic have remained limited. In this study, salivaricin A2 was purified and its covalent structure was characterized. N-terminal analogues of salivaricin A2 were generated to study the importance for bioactivity of the length and charge of the N-terminal amino acids. Analogue salivaricin A2(3-22) has no antibacterial activity and does not have an antagonistic effect on the native compound. The truncated analogue also lost its ability to bind to lipid II in a thin-layer chromatography (TLC) assay, suggesting that the N-terminal amino acids are important for binding to lipid II. The creation of N-terminal analogues of salivaricin A2 promoted a better understanding of the bioactivity of this antibiotic and further elucidated the structural importance of the N-terminal leader peptide. The antibacterial activity of salivaricin A2 is due not only to the presence of the positively charged N-terminal amino acid residues, but to the length of the N-terminal linear peptide. IMPORTANCE The amino acid composition of the N-terminal linear peptide of salivaricin A2 is crucial for function. Our study shows that the length of the amino acid residues in the linear peptide is crucial for salivaricin A2 antimicrobial activity. Very few type AII lantibiotic covalent structures have been confirmed. The characterization of the covalent structure of salivaricin A2 provides additional support for the predicted lanthionine and methyl-lanthionine ring formations present in this structural class of lantibiotics. Removal of the N-terminal Lys1 and Arg2 residues from the peptide causes a dramatic shift in the chemical shift values of amino acid residues 7 through 9, suggesting that the N-terminal amino acids contribute to a distinct structural conformer for the linear peptide region. The demonstration that the bioactivity could be partially restored with the substitution of N-terminal alanine residues supports further studies aimed at determining whether new analogues of salivaricin A2 for novel applications can be synthesized., (Copyright © 2018 American Society for Microbiology.)

Introduction: Lanthipeptides are a class of ribosomally synthesized and post-translationally modified peptides. Lanthipeptides with antimicrobial activity are referred to as lantibiotics. Lantibiotics are generally active against Gram-positive bacteria. However, some modifications have expanded their activity toward Gram-negative bacteria. Furthermore, additional functions aside from antibacterial activities have been reported for lanthipeptides. Areas covered: This review provides a synopsis of current anthipeptide research for potential therapeutics. The review highlights the current tools used for identifying lanthipeptides from genomic sequencing data. It also describes the current approaches that have been used to overcome the limitations in the purification and isolation of lanthipeptides. The status of lanthipeptides in terms of potential applications and approaches that are currently being done to promote the development of lanthipeptides as novel therapeutics are also discussed. Expert opinion: Significant improvements have been made to promote the discovery of new lanthipeptides, while, simultaneously, tools have been developed to promote their production and isolation. Lanthipeptides are showing significant promise for treating bacterial infections, as well as for new applications as anticancer and antiviral agents, or as a novel treatment for pain management. At the current rate of lanthipeptide discovery and isolation of the products, it is likely several new applications will be discovered.