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There is increasing evidence for a daily rhythm of µ-opioid receptor (MOR) efficacy and the development of alcohol dependence. Previous studies show that β-arrestin 2 (bArr2) has an impact on alcohol intake, at least partially mediated via modulation of MOR signaling, which in turn mediates the alcohol rewarding effects. Considering the interplay of circadian rhythms on MOR and alcohol dependence, we aimed to investigate bArr2 in alcohol dependence at different time points of the day/light cycle on the level of bArr2 mRNA (in situ hybridization), MOR availability (receptor autoradiography), and MOR signaling (Damgo-stimulated G-protein coupling) in the nucleus accumbens of alcohol-dependent and non-dependent Wistar rats. Using a microarray data set we found that bArr2, but not bArr1, shows a diurnal transcription pattern in the accumbens of naïve rats with higher expression levels during the active cycle. In 3-week abstinent rats, bArr2 is up-regulated in the accumbens at the beginning of the active cycle (ZT15), whereas no differences were found at the beginning of the inactive cycle (ZT3) compared with controls. This effect was accompanied by a specific down-regulation of MOR binding in the active cycle. Additionally, we detect a higher receptor coupling during the inactive cycle compared with the active cycle in alcohol-dependent animals. Together, we report daily rhythmicity for bArr2 expression linked to an inverse pattern of MOR, suggesting an involvement for bArr2 on circadian regulation of G-protein coupled receptors in alcohol dependence. The presented data may have implications for the development of novel bArr2-related treatment targets for alcoholism. (Figure presented.).

Background: COVID-19 is a worldwide threat because of the incessant spread of SARS-CoV-2 which urges the development of suitable antiviral drug to secure our society. Already, a group of peptides have been recommended for SARS-CoV-2, but not yet established. SARS-CoV-2 is an enveloped virus with hydrophobic fusion protein and spike glycoproteins. Methods: Here, we have summarized several reported amphiphilic peptides and their in-silico docking analysis with spike glycoprotein of SARS-CoV-2. Result: The result revealed the complex formation of spike protein and amphiphilic peptides with higher binding affinity. It was also observed that PalL1 (ARLPRTMVHPKPAQP), 10AN1 (FWFTLIKTQAKQPARYRRFC), THETA defensin (RCICGRGICRLL) and mucroporin M1 (LFRLIKSLIKRLVSAFK) showed the binding free energy more than -1000 kcal/mol. Molecular pI and hydrophobicity are also important factors of peptides to enhance the binding affinity with spike protein of SARS-CoV-2 Conclusion: In the light of these findings, it is necessary to check the real efficacy of amphiphilic peptides in-vitro to in-vivo experimental set up to develop an effective anti-SARS-CoV-2 peptide drug, which might help to control the current pandemic situation.

Background: COVID-19 is a life-threatening novel corona viral infection to our civilization and spreading rapidly. Tremendousefforts have been made by the researchers to search for a drug to control SARS-CoV-2. Methods: Here, a series of arsenical derivatives were optimized and analyzed with in silico study to search the inhibitor of RNA dependent RNA polymerase (RdRp), the major replication factor of SARS-CoV-2. All the optimized derivatives were blindly docked with RdRp of SARS-CoV-2 using iGEMDOCK v2.1. Results: Based on the lower idock score in the catalytic pocket of RdRp, darinaparsin (-82.52 kcal/- mol) was revealed to be the most effective among them. Darinaparsin strongly binds with both Nsp9 replicase protein (-8.77 kcal/mol) and Nsp15 endoribonuclease (-8.3 kcal/mol) of SARS-- CoV-2 as confirmed from the AutoDock analysis. During infection, the ssRNA of SARS-CoV-2 is translated into large polyproteins forming viral replication complex by specific proteases like 3CL protease and papain protease. This is also another target to control the virus infection where darinaparsin also performs the inhibitory role to proteases of 3CL protease (-7.69 kcal/mol) and papain protease (-8.43 kcal/mol). Conclusion: In the host cell, the furin protease serves as a gateway to the viral entry and darinaparsin docked with furin protease, which revealed a strong binding affinity. Thus, screening of potential arsenic drugs would help in providing the fast in-vitro to in-vivo analysis towards the development of therapeutics against SARS-CoV-2.

Infectious diseases caused by viruses have become a serious public health issue in the recent past, including the current pandemic situation of COVID-19. Enveloped viruses are most commonly known to cause emerging and recurring infectious diseases. Viral and cell membrane fusion is the major key event in the case of enveloped viruses that is required for their entry into the cell. Viral fusion proteins play an important role in the fusion process and in infection establishment. Because of this, the fusion process targeting antivirals become an interest to fight against viral diseases caused by the enveloped virus. Lower respiratory tract infections casing viruses like influenza, respiratory syncytial virus (RSV), and severe acute respiratory syndrome coronavirus (SARS-CoV) are examples of such enveloped viruses that are at the top in public health issues. Here, we summarized the viral fusion protein targeted antiviral peptides along with their mechanism and specific design to combat the viral fusion process. The pandemic COVID-19, severe respiratory syndrome disease is an outbreak worldwide. There are no definitive drugs yet, but few are in on-going trials. Here, an approach of fragmentbased drug design (FBDD) methodology is used to identify the broad spectrum agent target to the conserved region of fusion protein of SARS CoV-2. Three dipeptides (DL, LQ and ID) were chosen from the library and designed by the systematic combination along with their possible modifications of amino acids to the target sites. Designed peptides were docked with targeted fusion protein after energy minimization. Results show strong and significant binding affinity (DL = -60.1 kcal/mol; LQ = - 62.8 kcal/mol; ID= -71.5 kcal/mol) during interaction. Anyone of the active peptides from the developed libraries may help to block the target sites competitively to successfully control COVID-19.

Antheraea mylitta, a tropical non-mulberry silkworm, is cultivated for tasar silk production in India. Several defense molecules including few antimicrobial peptides (AMPs) and proteins have been identified from this insect. Here, we have isolated and purified an antimicrobial tri-peptide by sequential chromatographic separation procedures. The amino acid sequence of the peptide was determined as NH2-Gln-Ala-Lys-COOH (QAK) using MALDI MS/MS fragmentation analysis. Further, the peptide was synthesized in vitro following solid phase chemistry of peptide synthesis and acetylated by acetic anhydride reaction. Antimicrobial activities of non-acetylated and acetylated QAK were tested against both Escherichia coli and Staphylococcus aureus bacteria. Acetylated peptide inhibited bacterial growth more effectively and its minimum inhibitory concentration (MICs) was found lower than non-acetylated peptide. SEM studies revealed more membrane damage and release of intracellular materials like β-galactosidase enzyme from acetylated peptide treated bacteria in comparison to non-acetylated QAK. At MIC, acetylated peptide did not show any significant hemolytic activity against rabbit erythrocytes. The results suggest that acetylated-QAK is a promising new antimicrobial peptide and can be used for therapeutic purpose.

Abtract Humoral defenses are the major components of insect innate immune system that include the production of several soluble effector molecules from fat body and hemocytes, and released in to the hemolymph upon microbial infection. Hemolymph was collected from the fungal immunized fifth instar larvae of tasar silkworm, Antheraea mylitta, extracted with a mixture of solvent (methanol/glacial acetic acid/water) and fractionated through RP-HPLC. Several fractions were collected, lyophilized and their antifungal activity was tested against Candida albicans. Only the fraction showing strong antifungal activity was further purified via gel filtration chromatography and the purity of active compound was confirmed by thin layer chromatography which showed only single spot after staining with ninhydrin. The molecular mass of this purified compound was determined by high resolution mass spectrometry as 531 Da and analysis of 1H and 13C NMR spectral data along with mass fragmentation pattern indicated the probable structure of the isolated compound as symmetric bis-decanoate derivative. Scanning electron microscopic study revealed that the compound degraded fungal cell wall leading to its lysis and may be the major target for its antifungal activity. These results indicate that presence of this compound in the hemolymph of A. mylitta provides defense against fungal infection.

Antimicrobial proteins (AMPs) are small, cationic proteins that exhibit activity against bacteria, viruses, parasites, fungi as well as boost host-specific innate immune responses. Insects produce these AMPs in the fat body and hemocytes, and release them into the hemolymph upon microbial infection. Hemolymph was collected from the bacterially immunized fifth instar larvae of tasar silkworm, Antheraea mylitta, and an AMP was purified by organic solvent extraction followed by size exclusion and reverse-phase high-pressure liquid chromatography. The purity of AMP was confirmed by thin-layer chromatography and sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis. The molecular mass was determined by matrix-assisted laser desorption ionization-time of flight mass spectrometry as 14 kDa, and hence designated as AmAMP14. Peptide mass fingerprinting of trypsin-digested AmAMP14 followed by de novo sequencing of one peptide fragment by tandem mass spectrometry analysis revealed the amino acid sequences as CTSPKQCLPPCK. No homology was found in the database search and indicates it as a novel AMP. The minimum inhibitory concentration of the purified AmAMP14 was determined against Escherichia coli, Staphylococcus aureus, and Candida albicans as 30, 60, and 30 µg/ml, respectively. Electron microscopic examination of the AmAMP14-treated cells revealed membrane damage and release of cytoplasmic contents. All these results suggest the production of a novel 14 kDa AMP in the hemolymph of A. mylitta to provide defense against microbial infection.

The work demonstrate screening of several arsenical compounds against RdRp of coronavirus. The study implies out of all arsenical compounds, darinaparsin shows its most effective results based on in silico docking analysis. This study also confirmed the significant interaction between the active site of viral replicase protein, endoribonuclease protein and different proteases with darinaparsin.

Cancer is the leading cause of deaths worldwide, though significant advances have occurred in its diagnosis and treatment. The development of resistance against chemotherapeutic agents, their side effects, and non-specific toxicity urge to screen for the novel anticancer agent. Hence, the development of novel anticancer agents with a new mechanism of action has become a major scientific challenge. Bacteria and bacterially produced bioactive compounds have recently emerged as a promising alternative for cancer therapeutics. Bacterial anticancer agents such as antibiotics, bacteriocins, non-ribosomal peptides, polyketides, toxins, etc. These are adopted different mechanisms of actions such as apoptosis, necrosis, reduced angiogenesis, inhibition of translation and splicing, and obstructing essential signaling pathways to kill cancer cells. Also, live tumor-targeting bacteria provided a unique therapeutic alternative for cancer treatment. This review summarizes the anticancer properties and mechanism of actions of the anticancer agents of bacterial origin and antitumor bacteria along with their possible future applications in cancer therapeutics.

The reactivity of indole based bis-propargyl ethers 4a-4g under Garratt-Braverman condition (KOBut in refluxing toluene) has been studied. Interestingly, these propargyl systems with one arm attached with substituted 3-indolyl derivatives leaving the other arm unsubstituted produced the 3,4-furan fused dihydrocarbazole derivatives 6a-6g (and not the expected carbazole derivatives) as the predominant product (70–82%) making this methodology to access such derivatives an attractive route. The results are supported by computational studies and some of the carbazole derivatives showed good antifungal activities.

Long-term treatment with several conventional antibiotics can cause harmful side effects that can be alleviated by antioxidant therapy. Phenolic compounds (PCs) are the best source of antioxidants, and to identify the most suitable polyphenols for use as a supportive supplement during antibiotic therapy, this study screened a series of PCs to establish their antibacterial potential, including their biofilm and β-lactamase inhibition activity. Several PCs were tested for antibacterial activity against Staphylococcus epidermidis and Pseudomonas aeruginosa. Among them, tannic acid, epigallocatechin gallate, rutin, and eugenol showed the highest antibacterial activity. Epigallocatechin gallate, tannic acid, quercetin, and epicatechin inhibited a significant level of β-lactamase activity. Tannic acid and epigallocatechin gallate presented the highest β-lactamase inhibition potential in both in vitro and in silico analysis. In the present work, these two PCs were the most active agents, presenting both antibacterial activity and β-lactamase and biofilm inhibition ability.

Antimicrobial peptides (AMPs) are produced in all living organisms including insects in a non-specific manner, and act as innate immune defense arsenal against the invading pathogens. Muga silkworm (Antheraea assamensis) larvae were injected with Candida albicans and AMPs were isolated from the hemolymph after extracting with methanol, acetic acid and water mixture (90:1:9) and evaluated for antimicrobial activity against fungal and bacterial pathogens. Further purification was done through successive semipreparative and analytical reversed phase HPLC using C-18 column. The obtained fractions were collected, lyophilized and tested for antimicrobial activity. Among the HPLC fractions, one showed highest activity with MIC value of 64 µg/ml against Gram-negative bacteria, Escherichia coli and Enterobacter cloacae. Purity of this isolated peptide was confirmed by SDS-PAGE and TLC, and its molecular mass was determined as 9.052 kDa by MALDI-TOF mass spectrometry. From the mass fingerprinting analysis of this peptide after trypsin digestion a peptide fragment with molecular mass of 2622.7 Da was obtained. De novo sequencing of this peptide fragment following MS/MS analysis identified few amino acid residues as “KSGGGGWGS” with a total score of 46.9 with gloverin peptide of A. mylitta. The peptide inhibited biofilm formation of the Gram-negative bacterial pathogens. SEM study revealed that peptide disrupted bacterial cell wall to leach out intracellular materials and may be the major target for its antimicrobial activity.

The fluoroquinolones (FQs) are one the most successful class of synthetic antibiotics that primarily target the type II topoisomerases. With a pursuit to evaluate their genotoxicity, the present work established moderate to good DNA-damaging properties of some of the well-known and clinically prescribed fluoroquinolone antibiotics (2nd and 3rd generation). Hypochromic shift in UV–Vis absorption titration, fluorescence quenching in competitive ethidium bromide displacement assay (with calf-thymus DNA) and in-silico studies established DNA-intercalation with binding constants of the order 104. A basic Structure Activity Relationship (SAR) has been derived from the docking results. MTT assay has been also done to evaluate the effect of these antibiotics on cell viability. The expression level of specific DNA-glycosylase enzymes responsible for repairing the oxidized DNA bases are quantified through western blot analysis. The studies revealed that fluoroquinolone antibiotics initiate the genotoxic effect at a concentration of above 50 μg/mL. Recruitment of APE1 and NEIL1 was found to be significantly increased to remove the oxidized nucleobases.

Antimicrobial resistance (AMR) reported to increase globally at alarming levels in the recent past. A number of potential alternative solutions discussed and implemented to control AMR in bacterial pathogens. Stringent control over the clinical application of antibiotics for a reduction in uses is a special consideration along with alternative solutions to fight against AMR. Although alternatives to conventional antibiotics like antimicrobial peptides (AMP) might warrant serious consideration to fight against AMR, there is a thriving recognition for vaccines in encountering the problem of AMR. Vaccines can reduce the prevalence of AMR by reducing the number of specific pathogens, which result in cutting down the antimicrobial need and uses. However, conventional vaccines produced using live or attenuated microorganisms while the presence of immunologically redundant biological components or impurities might cause major side effects and health related problems. Here we discussed AMPs based vaccination strategies as an emerging concept to overcome the disadvantages of traditional vaccines while boosting the AMPs to control multidrug resistant bacteria or AMR. Nevertheless, the poor immune response is a major challenge in the case of peptide vaccines as minimal antigenic epitopes used for immunization in peptide vaccines.

Multi-drug resistance (MDR) bacteria pose a significant threat to our ability to effectively treat infections due to the development of several antibiotic resistant mechanisms. A major component in the development of the MDR phenotype in MDR bacteria is over expression of different-type of efflux pumps, which actively pump out antibacterial agents and biocides from the periplasm to the outside of the cell. Consequently, bacterial efflux pumps are an important target for developing novel antibacterial treatments. Potent efflux pump inhibitors (EPIs) could be used as adjunctive therapies that would increase the potency of existing antibiotics and decrease the emergence of MDR bacteria. Several potent inhibitors of efflux pumps have been reported which has been summarized here. All the natural and synthetic EPIs were optimized with Gaussian and Avogadro software. The optimized structures were docked with each class of efflux pumps and their bonding parameters were computed. The theoretical analyses were performed with density functional theory (DFT). Overall, computational study revealed a good trend of electrophilicity and ionization potential of the EPIs, the obtained average values are within in the range of 0.001414 AU ± 0.00032 and 0.208821 AU ± 0.015545, respectively. Interestingly, cathinone interacts with most of the efflux pumps among the tested inhibitors. The electrophilicity and ionization potential of cathinone are 0.00198 and 0.2388 AU, respectively. The study opens a new road for designing future-generation target-specific efflux pump inhibitors, as well as one molecule with multiple inhibition abilities.

While antibiotics remain as a major therapy against life threatening pathogenic infections, they often lead to side effects like rashes, gastrointestinal and central nervous system reactions to serious allergies or organ damage. These adverse effects alongside the emergence of multi-antibiotic resistant bacteria and the decline in the development of new antibiotics, have posed a serious impediment for effective antibiotic therapy. A paradigm shift in attitudes has led us to think about the possibility of controlling infections with the indigenous antimicrobial peptides synthesized by human beings. It has been observed that few transcription factors can stimulate more than three dozen defense peptides in the human system. Hence, during the infection stage, if we can induce these common factors, most of the infections could be healed from inside without the administration of any antibiotics. The efficiency of such peptides is being proven in clinical tests leading to the development of drugs.

Succumbing to Multi-Drug Resistant (MDR) bacteria is a great distress to the recent health care system. Out of the several attempts that have been made to kill MDR pathogens, a few gained short-lived success. The failures, of the discovered or innovated antimicrobials, were mostly due to their high level of toxicity to hosts and the phenomenal rate of developing resistance by the pathogens against the new arsenal. Recently, a few quantum dots were tested against the pathogenic bacteria and therefore, justified for potential stockpiling of next-generation antibacterial agents. The key players for antimicrobial properties of quantum dots are considered to be Reactive Oxygen Species (ROS). The mechanism of reaction between bacteria and quantum dots needs to be better understood. They are generally targeted towards the cell wall and membrane components as lipoteichoic acid and phosphatidyl glycerol of bacteria have been documented here. In this paper, we have attempted to simulate ZnS quantum dots and have analysed their mechanism of reaction as well as binding potential to the above bacterial membrane components using CDOCKER. Results have shown a high level of antibacterial activity towards several pathogenic bacteria which specify their potentiality for future generation antibacterial drug development.

This study aimed to explore the bactericidal activity of a feather-degraded active peptide against multiple-antibiotic-resistant (MAR) Staphylococcus aureus. An antibacterial peptide (ABP) was isolated from the chicken feathers containing fermented media of Paenibacillus woosongensis TKB2, a keratinolytic soil isolate. It was purified by HPLC, and its mass was found to be 4666.87 Da using matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) spectroscopy. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) values of this peptide were 22.5 and 90 μg/ml, respectively. SEM study revealed the distorted cell wall of the test strain along with pore formation. The possible reason for bactericidal activity of the peptide is due to generation of reactive oxygen species (ROS), resulting in membrane damage and leakage of intracellular protein. Complete sequence of the peptide was predicted and retrieved from the sequence database of chicken feather keratin after in silico trypsin digestion using ExPASy tools. Further, net charge, hydrophobicity (77.7 %) and molecular modelling of the peptide were evaluated for better understanding of its mode of action. The hydrophobic region (17 to 27) of the peptide may facilitate for initial attachment on the bacterial membrane. The ABP exhibited no adverse effects on RBC membrane and HT-29 human cell line. This cytosafe peptide can be exploited as an effective therapeutic agent to combat Staphylococcal infections.

Lignin, one of the most abundant renewable feedstock, is used to develop a biocompatible hydrogel as anti-infective ointment. A hydrophilic polyoxazoline chain is grafted through ring opening polymerization, possess homogeneous spherical nanoparticles of 10–15 nm. The copolymer was covalently modified with triazole moiety to fortify the antimicrobial and antibiofilm activities. The hydrogel was capable of down regulating the expression level of IL-1β in LPS induced macrophage cells, and to cause significant reduction of iNOS production. It supported cellular anti-inflammatory activity which was confirmed with luciferase assay, western blot, and NF-κB analysis. This novel lignin-based hydrogel tested in-vivo has shown the abilities to prevent infection of burn wound, aid healing, and an anti-inflammatory dressing material. The hydrogel reported here provides a new material platform to introduce a cost-effective and efficient ointment option after undertaking further work to look at its use in the area of clinical practice.

Cardanol is a non-isoprenoic phenolic lipid-mixture of distilled cashew nut shell liquid obtained from Anacardium occidentale. Herein, cardanol is purified from cashew nut shell liquid (CNSL) and synthesized to new compounds with different azo amphiphiles. These synthesized compounds are allowed to self-assembled in hydrophobic environment and checked antifungal activity against Candida albicans. Self-assembled structure of CABA showed higher antifungal activity (16 μg/mL) and chitin-binding ability in comparison to CAP and CANB. Furthermore, the self-assembled azo amphiphiles are immobilized with silver ions to prepare hydrogel which showed eight folds enhanced antifungal activity. Toxicity is reduced by several folds of self-assembled or hydrogel structure in comparison to pure compounds. Thus, the self-assembled structure of amphiphiles and their hydrogels have been found to be new macromolecules of interest with potential use as antifungal drugs.

A novel antibacterial ointment using bacitracin, specific for Gram-positive bacteria, and gramicidin S, a highly toxic antibacterial peptide, was here developed showing broad-spectrum antibacterial activities against pathogenic strains with less toxicity after self-assembly into nanofiber structures. Such structures were confirmed with scanning electron microscopy and CD analyses. In addition, in silico studies using docking associated with molecular dynamics were carried out to obtain information about fiber structural oligomerization. Thus, the bacitracin and gramicidin S-based self-assembled nanopeptide ribbon may be a successful ointment formulation for bacterial infection control.

Pharmaceutical industries have renewed interest in screening multifunctional bioactive peptides as a marketable product in health care applications. In this context, several animal and plant peptides with potential bioactivity have been reported. Milk proteins and peptides have received much attention as a source of health-enhancing components to be incorporated into nutraceuticals and functional foods. By using this source, 24 peptides have been fractionated and purified from human milk using RP-HPLC. Multifunctional roles including antimicrobial, antioxidant and growth stimulating activity have been evaluated in all 24 fractions. Nevertheless, only four fractions show multiple combined activities among them. Using a proteomic approach, two of these four peptides have been identified as lactoferrin derived peptide and kappa casein short chain peptide. Lactoferrin derived peptide (f8) is arginine-rich and kappa casein derived (f12) peptide is proline-rich. Both peptides (f8 and f12) showed antimicrobial activities against both Gram-positive and Gram-negative bacteria. Fraction 8 (f8) exhibits growth stimulating activity in 3T3 cell line and f12 shows higher free radical scavenging activity in comparison to other fractions. Finally, both peptides were in silico evaluated and some insights into their mechanism of action were provided. Thus, results indicate that these identified peptides have multiple biological activities which are valuable for the quick development of the neonate and may be considered as potential biotechnological products for nutraceutical industry.

Small cyclic peptides exhibiting potent biological activity have great potential for anticancer therapy. An antiproliferative cyclic octapeptide, cyclosaplin was purified from somatic seedlings of Santalum album L. (sandalwood) using gel filtration and RP-HPLC separation process. The molecular mass of purified peptide was found to be 858 Da and the sequence was determined by MALDI-ToF-PSD-MS as 'RLGDGCTR' (cyclic). The cytotoxic activity of the peptide was tested against human breast cancer (MDA-MB-231) cell line in a dose and time-dependent manner. The purified peptide exhibited significant antiproliferative activity with an IC50 2.06 μg/mL. In a mechanistic approach, apoptosis was observed in differential microscopic studies for peptide treated MDA-MB-231 cells, which was further confirmed by mitochondrial membrane potential, DNA fragmentation assay, cell cycle analysis and caspase 3 activities. The modeling and docking experiments revealed strong affinity (kcal/mol) of peptide toward EGFR and procaspase 3. The co-localization studies revealed that the peptide sensitizes MDA-MB-231 cells by possibly binding to EGFR and induces apoptosis. This unique cyclic octapeptide revealed to be a favorable candidate for development of anticancer agents.

Antipsychotic (AP) drugs have the potential to cause prolongation of the QT interval corrected for heart rate (QTc). As this risk is dose-dependent, it may be associated with the number of AP drugs concurrently prescribed, which is known to be associated with increased cumulative equivalent AP dosage. This study analysed whether AP dose mediates the relationship between polypharmacy and QTc interval. We used data from a cross-sectional survey that investigated the prevalence of QTc lengthening among people with psychiatric illnesses in Italy. AP polypharmacy was tested for evidence of association with AP dose and QTc interval using the Baron and Kenny mediational model. A total of 725 patients were included in this analysis. Of these, 186 (26%) were treated with two or more AP drugs (AP polypharmacy). The mean cumulative AP dose was significantly higher in those receiving AP polypharmacy (prescribed daily dose/defined daily dose = 2.93, standard deviation 1.31) than monotherapy (prescribed daily dose/defined daily dose = 0.82, standard deviation 0.77) (z = -12.62, p < 0.001). Similarly, the mean QTc interval was significantly longer in those receiving AP polypharmacy (mean = 420.86 milliseconds, standard deviation 27.16) than monotherapy (mean = 413.42 milliseconds, standard deviation 31.54) (z = -2.70, p = 0.006). The Baron and Kenny mediational analysis showed that, after adjustment for confounding variables, AP dose mediates the association between polypharmacy and QTc interval. The present study found that AP polypharmacy is associated with QTc interval, and this effect is mediated by AP dose. Given the high prevalence of AP polypharmacy in real-world clinical practice, clinicians should consider not only the myriad risk factors for QTc prolongation in their patients, but also that adding a second AP drug may further increase risk as compared with monotherapy.

Nowadays, peptide based disease prevention is an important topic in biomedical science, which may radically change the traditional use of biomaterials and improve the life quality. Self-assembled nanostructured peptides have been receiving extreme attention in the drug delivery field due their high biocompatibility levels, better loading capacity, extended circulation and localization in required target site. This article focuses on the composition and synthesis of different forms of self-assembled peptide nanostructures as nanotubes, nanofibers, nanoparticles, nanotapes and nanogels. The most important properties for their self assembled mechanism and their biomedical applications are also discussed. Various potential applications of nanostructures peptide could be developed designed for therapeutic agent's delivery, biosensors, anticancerous and antimicrobial activities.

Phytopathogens cause economic losses in agribusiness. Plant-derived compounds have been proposed to overcome this problem, including the antimicrobial peptides (AMPs). This paper reports the identification of Ps-AFP1, a novel AMP isolated from the Pisum sativum radicle. Ps-AFP1 was purified and evaluated against phytopathogenic fungi, showing clear effectiveness. In silico analyses were performed, suggesting an unusual fold and disulfide bond pattern. A novel fold and a novel AMP class were here proposed, the αβ-trumpet fold and αβ-trumpet peptides, respectively. The name αβ-trumpet was created due to the peptide's fold, which resembles the musical instrument. The Ps-AFP1 mechanism of action was also proposed. Microscopic analyses revealed that Ps-AFP1 could affect the fungus during the hyphal elongation from spore germination. Furthermore, confocal microscopy performed with Ps-AFP1 labeled with FITC shows that the peptide was localized at high concentration along the fungal cell surface. Due to low cellular disruption rates, it seems that the main target is the fungal cell wall. The binding thermogram and isothermal titration, molecular dynamics and docking analyses were also performed, showing that Ps-AFP1 could bind to chitin producing a stable complex. Data here reported provided novel structural–functional insights into the αβ-trumpet peptide fold.

Towards the search for a new generation of antibiotics to control methicillin-resistant Staphylococcus aureus (MRSA), the design and synthesis of various bis indolyl methane (BIM) derivatives based on their different electron donor and acceptor properties of the substituents have been made, in which boronic acid derivatives of BIM are found to be active against MRSA. The observed evidence with the lead compound reveals their strong anti-MRSA activity, which paves the way of design and further development of a new generation antibiotics.

This work is licensed under a Creative Commons Attribution 4.0 International License. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in the credit line; if the material is not included under the Creative Commons license, users will need to obtain permission from the license holder to reproduce the material. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/, Clavanins is a class of peptides (23aa) histidine-rich, free of post-translational modifications. Clavanins have been studied largely for their ability to disrupt bacterial membranes. In the present study, the interaction of clavanin A with membranes was assessed by dynamic light scattering, zeta potential and permeabilization assays. We observed through those assays that clavanin A lysis bacterial cells at concentrations corresponding to its MIC. Further, the structure and function of clavanin A was investigated. To better understand how clavanin interacted with bacteria, its NMR structure was elucidated. The solution state NMR structure of clavanin A in the presence of TFE-d3 indicated an α-helical conformation. Secondary structures, based on circular dichroism measurements in anionic sodium dodecyl sulfate (SDS) and TFE (2,2,2-trifluorethanol), in silico lipid-peptide docking and molecular simulations with lipids DPPC and DOPC revealed that clavanin A can adopt a variety of folds, possibly influencing its different functions. Microcalorimetry assays revealed that clavanin A was capable of discriminating between different lipids. Finally, clavanin A was found to eradicate bacterial biofilms representing a previously unrecognized function., We would like to thank CNPq, CAPES (Ciências sem Fronteiras), FAPDF and FUNDECT. D.G. acknowledges Fundação para a Ciência e a Tecnologia - Ministério da Educação e Ciência (FCT-MEC, Portugal) for fellowship SFRH/BPD/73500/2010 and A.S.V. for funding within the FCT Investigator Programme (IF/00803/2012).

Probiotics are unique bacteria that offer several therapeutic benefits to human beings when administered in optimum amounts. Probiotics are able to produce antimicrobial substances, which stimulate the body's immune responses. Here, we review in detail the anti-infective peptides derived from probiotics and their potential immunomodulatory and anti-inflammatory activities, including a major role in cross-talk between probiotics and gut microbiota under adverse conditions. Insights from the engineered cell surface of probiotics may provide novel anti-infective therapy by heterologous expression of receptor peptides of bacterial toxins. It may be possible to use antigenic peptides from viral pathogens as live vaccines. Another possibility is to generate antiviral peptides that bind directly to virus particles, while some peptides exert anti-inflammatory and anticancer effects. Some extracellular polymeric substances might serve as anti-infective peptides. These avenues of treatment have remained largely unexplored to date, despite their potential in generating powerful anti-inflammatory and anti-infective products.

An extracellular antifungal protein of 28 kDa (exAFP-C28) was identified from an endophytic fungus Colletotrichum sp. DM-06. After purification, the MIC value of exAFP-C28 against Candida albicans, a well-known human pathogenic fungus was found to be 32 μg/mL that unaffected the human red blood cells. The antifungal activity associated with exAFP-C28 was manifested by the increased membrane permeability of C. albicans cells followed by disruption. Proteomics and bioinformatics analyses revealed that several peptide fragments of exAFP-C28 have identity with the bacterial 50S ribosomal protein L10, and a stretch of 55 amino acids of two peptide fragments corresponding to the Nterminus of L10 protein is capable of forming amphipathic helix required for membrane penetration. Taken together, our results suggest that the exAFP-C28 protein from Colletotrichum sp. DM-06 is a promising therapeutic agent in controlling candidiasis disease in animals including humans.

Aims To examine tannic acid (TA) utilization capacity by nitrogen-fixing bacteria, Azotobacter sp. SSB81, and identify the intermediate products during biotransformation. Another aim of this work is to investigate the effects of TA on major biopolymers like extracellular polysaccharide (EPS) and polyhydroxybutyrate (PHB) synthesis. Methods and results Tannic acid utilization and tolerance capacity of the strain was determined according to CLSI method. Intermediate products were identified using high-performance liquid chromatography, LC-MS/MS and (1) H NMR analysis. Intermediates were quantified by multiple reactions monitoring using LC-MS/MS. The strain was able to tolerate a high level of TA and utilized through enzymatic system. Growth of Azotobacter in TA-supplemented medium was characterized by an extended lag phase and decreased growth rate. Presence of TA catalytic enzymes as tannase, polyphenol oxidase (PPO) and phenol decarboxylase was confirmed in cell lysate using their specific substrates. PPO activity was more prominent in TA-supplemented mineral medium after 48 h of growth when gallic to ellagic acid (EA) reversible reaction was remarkable. Phase contrast and scanning electron microscopic analysis revealed elongated and irregular size of Azotobacter cells in response to TA. (1) H NMR analysis indicated that TA was transformed into gallic acid (GA), EA and pyrogallol. Biopolymer (EPS and PHB) production was decreased several folds in the presence of TA compared with cells grown in only glucose medium. Conclusions This is the first evidence on the biotransformation of TA by Azotobacter and also elevated level of EA production from gallotannins. Azotobacter has developed the mechanism to utilize TA for their carbon and energy source. Significance and impact of the study The widespread occurrence and exploitation of Azotobacter sp. strain SSB81 in agricultural and forest soil have an additional advantage to utilize the soil-accumulated TA and detoxifies the allelopathic effect of constant accumulated TA in soil.

The increasing threats of multidrug resistant fungal pathogens, several studies have been focused to identify novel antifungal plant peptides with unique characteristics. Plants have been defending themselves against fungal infection by generating effective antifungal molecules. Here, a novel antifungal peptide with molecular mass of 4.0 kDa was purified from the pericarp of D. stramonium using reversed phase chromatography system after acetic acid extraction. Presence of sugar moieties (-GlcNAc-) in the peptide have been confirmed using thin layer chromatographic (TLC), CD polarimeter, and MALDI MS analysis. Complete amino acid sequences of this peptide by PSD MALDI MS analysis revealed to contain hydroxyproline in the centre of two cysteine residues. After sequencing "TFPKCAPTRhyPhy PGPKhyPCDINNFKSKFWHIWRA-(GlcNAc-)Asn" peptide named as "datucin." Antifungal sensitivity of datucin have been performed for both planktonic cells and biofilm phenotype ofa multidrug resistant clinical isolates, C. albicans and showed a MIC and MBEC values of 1 microM and 2 microM, respectively. Hence, D. stramonium offers a potential source of novel antifungal peptide datucin with possible utility in antifungal chemotherapy.

The repair of reactive oxygen species-induced base lesions and single strand breaks (SSBs) in the nuclear genome via the base excision (BER) and SSB repair (SSBR) pathways, respectively, is well characterize, and important for maintaining genomic integrity. However, the role of mitochondrial (mt) BER and SSBR proteins in mt genome maintenance is not completely clear. Here we show the presence of the oxidized base-specific DNA glycosylase Nei-like 2 (NEIL2) and the DNA end-processing enzyme polynucleotide kinase 3′-phosphatase (PNKP) in purified human mitochondrial extracts (MEs). Confocal microscopy revealed co-localization of PNKP and NEIL2 with the mitochondrion-specific protein cytochrome c oxidase subunit 2 (MT-CO2). Further, chromatin immunoprecipitation analysis showed association of NEIL2 and PNKP with the mitochondrial genes MT-CO2 and MT-CO3 (cytochrome c oxidase subunit 3); importantly, both enzymes also associated with the mitochondrion-specific DNA polymerase γ. In cell association of NEIL2 and PNKP with polymerase γ was further confirmed by proximity ligation assays. PNKP-depleted ME showed a significant decrease in both BER and SSBR activities, and PNKP was found to be the major 3′-phosphatase in human ME. Furthermore, individual depletion of NEIL2 and PNKP in human HEK293 cells caused increased levels of oxidized bases and SSBs in the mt genome, respectively. Taken together, these studies demonstrate the critical role of NEIL2 and PNKP in maintenance of the mammalian mitochondrial genome.

The antimicrobial peptides (AMPs) are evolutionarily ancient molecules that act as components of the innate immune system. Recently, it was demonstrated that a single AMP can perform various functions; this ability is known as “peptide promiscuity.” However, little is known about promiscuity in plant AMPs without disulfide bonds. This study was carried out to evaluate the promiscuity of Cn-AMP1: a promising disulfide-free plant peptide with reduced size and cationic and hydrophobic properties. Its activity against human pathogenic bacteria and fungal pathogens, as well as its in vitro immunostimulatory activity and effects on cancerous and healthy mammalian cell proliferation were studied here. Cn-AMP1 exerts antimicrobial effects against Gram-positive bacteria, Gram-negative bacteria, and fungi. Moreover, tumor cell viability activity in Caco-2 cells, as well as immunostimulatory activity by evaluating upregulated inflammatory-cytokine secretion by monocytes was also positively observed. Cn-AMP1 does not exhibit a well-defined conformation in aqueous solution and probably undergoes a 310-helix transition in hydrophobic environments. The experimental results support the promiscuous activity of Cn-AMP1, presenting a wide range of activities, including antibacterial, antifungal, and immunostimulatory activity. In the future, Cn-AMP1 should be used in the development of novel biopharmaceuticals, mainly due to its reduced size and broad spectrum of activity. © 2012 Wiley Periodicals, Inc. Biopolymers (Pept Sci) 98: 322–331, 2012.

Due to recent emergence of fungal pathogens resistant to current antifungal therapies, several studies have been focused on screening of plant peptides to find novel compounds having antifungal activities. Here, a novel antifungal plant peptide, with molecular mass of 1230 Da was purified from fruits of Trapa natans by reverse phase high performance liquid chromatography using 300SB-C18 column and named as Tn -AFP1. Determination of complete amino acid sequences of this peptide by tandem mass spectrometry showed to contain following eleven amino acid residues: LMCTHPLDCSN. Purified Tn -AFP1 showed the inhibition of Candida tropicalis growth in vitro and disrupted the biofilm formation in a concentration dependent manner. It also showed downregulation of MDR1 and ERG11 gene expression in real time-PCR analysis. In silico molecular modeling predicted the structure of Tn -AFP1 as a single coil attached by a unique disulfide bond. Characterization of Tn -AFP1 could contribute in designing novel derivative(s) of this peptide for the development of more effective antimycotic compounds.

Increasing tendency of clinical bacterial strains resistant to conventional antibiotics has being a great challenge to the public's health. Antimicrobial peptides, a new class of antibiotics is known to have the activity against a wide range of bacteria resistant to conventional antibiotics. An antimicrobial peptide of 1676 Da was purified from Bellamya bengalensis, a fresh water snail, using ultrafiltration and reversed phase liquid chromatography. The effect of this peptide on Staphylococcus epidermidis resistant to ampicillin and chloramphenicol was investigated; the MIC and MBC values were 8 μg/ml and 16 μg/ml, respectively. Complete sequence of the peptide was determined by tandem mass spectrometry (MS/MS). Further, peptide net charge, hydrophobicity and molecular modeling were evaluated in silico for better understanding the probable mechanisms of action. The peptide showed the specificity to bacterial membranes. Hence, this reported peptide revealed a promising candidate to contribute in the development of therapeutic agent for Staphylococcal infections.

Preferential repair of bulky DNA adducts from the transcribed genes via nucleotide excision repair is well characterized in mammalian cells. However, definitive evidence is lacking for similar repair of oxidized bases, the major endogenous DNA lesions. Here we show that the oxidized base-specific human DNA glycosylase NEIL2 associates with RNA polymerase II and the transcriptional regulator heterogeneous nuclear ribonucleoprotein-U (hnRNP-U), both in vitro and in cells. NEIL2 immunocomplexes from cell extracts preferentially repaired the mutagenic cytosine oxidation product 5-hydroxyuracil in the transcribed strand. In a reconstituted system, we also observed NEIL2-initiated transcription-dependent base excision repair of 5-hydroxyuracil in the transcribed strand, with hnRNP-U playing a critical role. Chromatin immunoprecipitation/reimmunoprecipitation studies showed association of NEIL2, RNA polymerase II, and hnRNP-U on transcribed but not on transcriptionally silent genes. Furthermore, NEIL2-depleted cells accumulated more DNA damage in active than in silent genes. These results strongly support the preferential role of NEIL2 in repairing oxidized bases in the transcribed genes of mammalian cells.

A simple and rapid method for the identification and quantification of vitamin B(2) and B(12) in human urine has been developed using reverse phase high performance liquid chromatography (HPLC) and the peaks identity were confirmed by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS). HPLC separation was performed in single wavelength detector (lambda(365)) mode and separated isocratically using mobile phase methanol: 1mM aqueous TFA (1:4) in C18 column. The calibration graphs plotted with different concentrations of vitamin B(2) and B(12) was linear with a correlation coefficients (r(2))=0.9975 and 0.9985, respectively. The recoveries of vitamin B(2) and B(12) were above 87% and 90%, respectively. The results of this present study suggest that the proposed method may be simple and convenient way of identifying and quantifying vitamin B(2) and B(12) from human urine.

Infections caused by pathogenic bacteria could cause an expressive negative impact on human health. A significant enhance in resistance to commercial antibiotics has been observed in all kinds of pathogenic bacteria. In order to find novel approaches to control such common infections, a wide number of defense peptides with bactericidal properties have been characterized. In this report, three peptides lower than 3 kDa were purified and identified from green coconut ( Cocos nucifera L.) water by using reversed phase-high performance liquid chromatography (HPLC), showing molecular masses of 858 Da, 1249 Da and 950 Da. First one, named Cn -AMP1, was extremely efficient against both Gram-positive and Gram-negative bacteria, being MICs calculated for three peptides. All complete sequences were determined by MALDI-ToF analysis showing no identity in databanks. Moreover, peptide net charge and hydrophobicity of each peptide was in silico evaluated. Finally molecular modeling and dynamics were also applied generating peptides three-dimensional structures, indicating a better explanation to probable mechanisms of action. Cn -AMPs here reported show remarkable potential to contribute in the development of novel antibiotics from natural sources.

Phenolic acids are active antimicrobial compounds and root signaling molecules that play important roles in plant defense responses. They are generally present in plants as glycosides or esters. A range of soluble and bound phenolic acids were detected in roots and root nodules of Arachis hypogaea L., among which five were identified by high performance liquid chromatography (HPLC) coupled with UV–Vis diode array detector (DAD), viz., p-coumaric acid (p-com), p-hydroxybenzaldehyde (HBAld), p-hydroxybenzoic acid (HBA), caffeic acid (CA) and protocatechuic acid (PA). Para-coumaric acid was constitutively present in all fractions whereas HBA was present in the soluble form only in young nodules. CA and PA were mostly present in the wall bound fraction. The root nodules contain higher concentration of phenolic acids than non-nodulated roots and presence of peroxidase and polyphenol oxidase indicate the metabolism of phenolic acids in roots and root nodules. These results indicate that phenolic acids (p-com and CA) in bound-glycosidic or ester forms were major components in cell wall fortification which provide protection to the root nodule from pathogen attack.

An extracellular polysaccharide (EPS) was produced by a Rhizobium sp. isolated from the root nodules of Vigna mungo (L.) Hepper. Maximum EPS production (346 mg l(-1)) was when the yeast extract basal medium was supplemented with mannitol (1%), biotin (1.5 mg l(-1)) and asparagine (0.3%). Ribose (53%) and mannose (47%) were the principle monomers of the EPS. Chemical, chromatographic and spectroscopic analysis showed that this polymer, which has Man(4)Rib(1) as an oligomeric subunit, has an apparent molecular mass of 750 kDa.

A range of phenolic acids, viz., p-coumaric acid, 4-hydroxybenzaldehyde, 4-hydroxybenzoic acid, protocatechuic acid, caffeic acid, ferulic acid, and cinnamic acid have been isolated and identified by LC-MS analysis in the roots and root nodules of Mimosa pudica. The effects of identified phenolic acids on the regulation of nodulation (nod) genes have been evaluated in a betarhizobium isolate of M. pudica root nodule. Protocatechuic acid and p-hydroxybenzoic acid were most effective in inducing nod gene, whereas caffeic acid had no significant effect. Phenylalanine ammonia lyase, peroxidase, and polyphenol oxidase activities were estimated, indicating regulation and metabolism of phenolic acids in root nodules. These results showed that nodD gene expression of betarhizobium is regulated by simple phenolic acids such as protocatechuic acid and p-hydroxybenzoic acid present in host root nodule and sustains nodule organogenesis.

Amphotericin B and anidulafungin are widely used antifungal drugs for the treatment of systemic and serious mycoses. Amphotericin B is a relatively toxic drug which has long been established. This study is first of its kind to systematically investigate the nature of binding to DNA, and to evaluate intercalation of AMP-B or ANIDULA with the aid of UV-Vis, ITC, and CD spectroscopy. The binding affinity of AMP-B with exclusion sites of 4.68 base pairs (1.2 × 10(5) M(-1)) was found to be higher than that of ANIDULA with exclusion sites of 6.67 base pairs (3.78 × 10(4) M(-1)); consistent with the binding affinity values obtained for AMP-B (10(5) M(-1)) and ANIDULA (10(4) M(-1)). The binding of two drugs with double-stranded DNA was favoured by negative enthalpy as well as negative entropy changes. The intercalation of drugs to duplex polynucleotide induced changes in the intrinsic CD spectra and revealed comparatively higher affinity towards AMP-B than ANIDULA. Molecular docking studies revealed that the negative binding energy was higher in the case of AMP-B reflecting more affinity towards single-stranded DNA. The results of the cytotoxicity, immunoblotting, and gene specific LA-QPCR assay have indicated that ANIDULA is less genotoxic than AMP-B. Hence, the superiority of ANIDULA over AMP-B as a systemic antifungal drug has been established beyond doubt.

Antimicrobial peptides form part of the first line of defense against pathogens for many organisms. Current treatments for fungal infections are limited by drug toxicity and pathogen resistance. Cm-p5 (SRSELIVHQRLF), a peptide derived from the marine mollusk Cenchritis muricatus peptide Cm-p1, has a significantly increased fungistatic activity against pathogenic Candida albicans (minimal inhibitory concentration, 10 µg/ml; EC50, 1.146 µg/ml) while exhibiting low toxic effects against a cultured mammalian cell line. Cm-p5 as characterized by circular dichroism and nuclear magnetic resonance revealed an α-helical structure in membrane-mimetic conditions and a tendency to random coil folding in aqueous solutions. Additional studies modeling Cm-p5 binding to a phosphatidylserine bilayer in silico and isothermal titration calorimetry using lipid monophases demonstrated that Cm-p5 has a high affinity for the phospholipids of fungal membranes (phosphatidylserine and phosphatidylethanolamine), only moderate interactions with a mammalian membrane phospholipid, low interaction with ergosterol, and no interaction with chitin. Adhesion of Cm-p5 to living C. albicans cells was confirmed by fluorescence microscopy with FITC-labeled peptide. In a systemic candidiasis model in mice, intraperitoneal administration of Cm-p5 was unable to control the fungal kidney burden, although its low amphiphaticity could be modified to generate new derivatives with improved fungicidal activity and stability.—López-Abarrategui, C., McBeth, C., Mandal, S. M., Sun, Z. J., Heffron, G., Alba-Menéndez, A., Migliolo, L., Reyes-Acosta, O., García-Villarino, M., Nolasco, D. O., Falcão, R., Cherobim, M. D., Dias, S. C., Brandt, W., Wessjohann, L., Starnbach, M., Franco, O. L., Otero-González, A. J. Cm-p5: an antifungal hydrophilic peptide derived from the coastal mollusk Cenchritis muricatus (Gastropoda: Littorinidae).

Effects of glucose on the susceptibility of antifungal agents were investigated against Candida spp. Increasing the concentration of glucose decreased the activity of antifungal agents; voriconazole was the most affected drugs followed by amphotericin B. No significant change has been observed for anidulafungin. Biophysical interactions between antifungal agents with glucose molecules were investigated using isothermal titration calorimetry, Fourier transform infrared, and (1)H NMR. Glucose has a higher affinity to bind with voriconazole by hydrogen bonding and decrease the susceptibility of antifungal agents during chemotherapy. In addition to confirming the results observed in vitro, theoretical docking studies demonstrated that voriconazole presented three important hydrogen bonds and amphotericin B presented two hydrogen bonds that stabilized the glucose. In vivo results also suggest that the physiologically relevant higher glucose level in the bloodstream of diabetes mellitus mice might interact with the available selective agents during antifungal therapy, thus decreasing glucose activity by complex formation. Thus, proper selection of drugs for diabetes mellitus patients is important to control infectious diseases.

Multifunctional and promiscuous antimicrobial peptides (AMPs) can be used as an efficient strategy to control pathogens. However, little is known about the structural properties of plant promiscuous AMPs without disulfide bonds. CD and NMR were used to elucidate the structure of the promiscuous peptide Cn-AMP1, a disulfide-free peptide isolated from green coconut water. Data here reported shows that peptide structure is transitory and could be different according to the micro-environment. In this regard, Cn-AMP1 showed a random coil in a water environment and an α-helical structure in the presence of SDS-d25 micelles. Moreover, deuterium exchange experiments showed that Gly4, Arg5 and Met9 residues are less accessible to solvent, suggesting that flexibility and cationic charges seem to be essential for Cn-AMP1 multiple activities.

Plants produce a variety of proteins and peptides which are involved in their defense against pathogens. Serine protease inhibitors are a well-established class of inhibitors correlated with plant defense. Increased levels of protease inhibitors delay cell damage and expand the cell's life-span. Recently, the rapid emergence of antibiotic-resistant microbial pathogens has prompted immense interest in purifying novel antimicrobial proteins or peptides from plant sources. Usually, the purification of protease inhibitors is accomplished by salt-extraction, ultrafiltration and affinity chromatography. Here, we developed a novel approach based on iron oxide nanoparticles conjugated to dextran functionalized with trypsin beads that accelerate the quick screening and purification of antimicrobial peptides with serine protease inhibitor activity. The method described here also works for screening other inhibitors using particular protein kinases, and it is therefore a novel tool for use as the leading method in the development of novel antimicrobial agents with protease inhibitory activity. Finally, and no less important, molecular modelling and dynamics studies of a homologous inhibitor studied here with Escherichia coli trypsin and chymotrypsin are provided in order to shed some light on inhibitor–enzyme interactions.

An antifungal lipopeptide fengycin, producing strain SM1 was isolated from farm land soil sample and identified as Bacillus thuringienesis strain SM1 by using 16S rDNA analysis. Fengycin detected in the culture extract was further purified using HPLC and showed a molecular mass of 1492.8 Da by MALDI-TOF-MS analysis. Purified fengycin was allowed to construct their self-assembled structure onto a hydrophobic surface showing a clear improvement of antibacterial activity. In self-assembly, fengycin adapts a spherical micelle core shell like structure. Self-assembled fengycin may be a successful antimicrobial compound modifying its action from confined antifungal function. Besides it can open up a new area of research in supramolecullar lipopeptide based compound making. This can revealed the mode of action of this unique self-assembled structure to fully evaluate its potential for use as an antimicrobial drug to control the emergence of bacterial infection.