Cyclic Dipeptide Cyclo Research Articles

Cyclic di-peptide Cyclo (L-Phe-L-Pro) mitigates the quorum-sensing mediated virulence in Salmonella typhi and biofilm formation in poultry and plastic system

Salmonella typhi is a predominant foodborne pathogen, posing a significant threat to food quality and safety through typhoid outbreaks. Quorum sensing (QS) and its pathways are crucial for typhoid antigen regulation and S. typhi-mediated spoilage and pathogenesis. Therefore, this study intends to identify secondary metabolites from extracts of 15 bacterial isolates sourced from wild Mangifera indica (mango) brine pickles as novel anti-QS agents against S. typhi. Cell-free supernatants (CFS) of 15 bacterial isolates from fermented mango brine pickles were assessed and identified for anti-QS positive metabolites using GC-MS. Anti-QS metabolite was tested on early and mature biofilms, as well as violacein inhibition, EPS, cellular adherence, motility, antibiotics, and immune resistance of S. typhi. Gene expression and in-silico studies were also conducted to validate the compound's efficacy. The results depicted that among the 15 isolates, Bacillus subtilis P89 exhibited potent anti-QS properties. Gas chromatography-mass spectrometry (GC-MS) analysis identified twelve potential quorum-sensing inhibiting compounds, with Cyclo (L-Phenylalanine-L-Proline) (CPP) demonstrating the maximum anti-QS activity against S. typhi. CPP disrupted the pathogen's early and mature biofilms, reducing exo-polymeric substance production, cell hydrophobicity, and motility at sub-lethal concentrations. It also enhanced the immune response against S. typhi and improved traditional efficacy. Real-time qPCR analyses revealed CPP's modulation of essential quorum-sensing regulatory genes. Molecular docking studies indicated competitive binding of CPP with the SdiA receptor. CPP inhibited S. typhi biofilm formation on chicken skin, plastics, and eggshells. To our knowledge, this study is the first to highlight CPP's potential in food and clinical environments to combat S. typhi via quorum sensing-based mechanisms.

Does Chiral Sensitivity of a Structure Depend on the Metal Core? Alkali Ion Complexes of Cyclo(Tyr-Tyr).

Alkali metal complexes of cyclic dipeptide cyclo Tyr-Tyr have been studied under cryogenic ion trap conditions. Their structure was obtained by combining Infra-Red Photo-Dissociation (IRPD) and quantum chemical calculations. The structural motif strongly depends on the relative chirality of the tyrosine residues. For residues of identical chirality, the cation interacts with one amide oxygen and one of the aromatic rings only; the distance between the aromatic rings does not change with the nature of the metal. In contrast, for residues of opposite chirality, the metal cation is located in between the two aromatic rings and interacts with both of them. The distance between the two aromatic rings strongly depends on the metal. Electronic spectra obtained by Ultra Violet Photodissociation (UVPD) spectroscopy and analysis of the UV photo-fragments shed light on the excited state deactivation processes, which depend on both the chirality of the residue and that of the metal ion core. Na+ stands out by the presence of low-lying charge transfer states resulting in the broadening of the electronic spectrum.

A model for microbial interactions and metabolomic alterations in Candida glabrata-Staphylococcus epidermidis dual-species biofilms.

The fungus Candida glabrata and the bacterium Staphylococcus epidermidis are important biofilm-forming microorganisms responsible of nosocomial infections in patients. In addition to causing single-species disease, these microorganisms are also involved in polymicrobial infections leading to an increased antimicrobial resistance. To expand knowledge about polymicrobial biofilms, in this study we investigate the formation of single- and dual-species biofilms of these two opportunistic pathogens employing several complementary approaches. First, biofilm biomass, biofilm metabolic activity and the microbial composition in single- and dual-species biofilms were assessed and compared. Then, the expression of three genes of C. glabrata and three genes of S. epidermidis positively related to the process of biofilm formation was evaluated. Although S. epidermidis is a stronger biofilm producer than C. glabrata, both biological and genetic data indicate that S. epidermidis growth is inhibited by C. glabrata which dominates the dual-species biofilms. To better understand the mechanisms of the interactions between the two microorganisms, a broad GC-MS metabolomic dataset of extracellular metabolites for planktonic, single- and dual-species biofilm cultures of C. glabrata and S. epidermidis was collected. As demonstrated by Partial Least Squares Discriminant Analysis (PLS-DA) of GC-MS metabolomic data, planktonic cultures, single- and dual-species biofilms can be sharply differentiated from each other by the nature and levels of an assortment of primary and secondary metabolites secreted in the culture medium. However, according to our data, 2-phenylethanol (secreted by C. glabrata) and the synergistically combined antifungal activity of 3-phenyllactic acid and of the cyclic dipeptide cyclo-(l-Pro-l-Trp) (secreted by S. epidermidis) play a major role in the race of the two microorganisms for predominance and survival.

A novel cyclic dipeptide from Lactiplantibacillus plantarum MC39 inhibits proliferation of multidrug-resistant Klebsiella pneumoniae W8 and Enterobacter hormaechei U25

The present study pays special attention to lactic acid bacteria products, cyclic dipeptides (CDPs), as an effective prevention strategy that can be used to eradicate the threats posed by multidrug-resistant (MDR) Enterobacteriaceae. Herein, Lactiplantibacillus plantarum MC39, a strain isolated from an artisanal Egyptian cheese, was a producer of a novel CDP that could strongly inhibit, in vitro, MDR Klebsiella pneumoniae W8 and Enterobacter hormaechei U25, for the first time. Following partial purification with 80% ammonium sulphate precipitation and Sephadex G-100 chromatography, this biomolecule was identified as the cyclic dipeptide cyclo(Ala–Glu) using IR, HNMR, and LC–MS. Cyclo(Ala–Glu) was thermostable up to 121 °C, insensitive to pepsin and α-chymotrypsin, tolerant to organic solvents, active at acidic pH, and stable for 45 days of storage. Thus, it shows clear evidence to be usable as a bio-preservative in dairy products and as a potential therapeutic agent, controlling MDR K. pneumoniae and E. hormaechei.

Glycosylation increases the anti-QS as well as anti-biofilm and anti-adhesion ability of the cyclo (L-Trp-L-Ser) against Pseudomonas aeruginosa

Pseudomonas aeruginosa infections pose a huge threat to cystic fibrosis patients, as well as those suffering from immunodeficiency. Antimicrobial resistance, especially multi-drug resistance, due to its ability to aggregate the compact biofilm, makes it more inefficient to treat this pathogen with traditional antibiotics. Biofilm and quorum sensing (QS) have become the alternative targets for treating P. aeruginosa infections. Previously, a cyclic dipeptide cyclo(L-Trp-L-Ser) has been identified as a QS inhibitor of P. aeruginosa. On the other hand, some monosaccharides have been proved lectin-targeting behavior and to mediate biofilm formation and adhesion of P. aeruginosa. We constructed novel cyclic dipeptide-carbohydrate conjugates as a low molecular weight dual-functional QS inhibitor, which can not only enhance its anti-QS activity but also enable good anti-biofilm and anti-adhesion ability. The IC50 of galactosylated c(WS) on biofilm formation and glass adhesion was 1/6 and 1/4 of that of the unmodified cyclic dipeptide, respectively. And the ability to eliminate the preformed biofilm was increased 10-fold. Furthermore, the carbohydrate conjugates can increase the germicidal efficiency of clinical antibiotic azithromycin when used synergistically. Our results provide a novel scaffold for developing anti-virulence adjuvants when taken with clinical antibiotics.

S-Benzyl cysteine based cyclic dipeptide super hydrogelator: Enhancing efficacy of an anticancer drug via sustainable release.

Peptide-based low molecular weight supramolecular hydrogels hold promising aspects in various fields of application especially in biomaterial and biomedical sciences such as drug delivery, wound healing, tissue engineering, cell proliferation, and so on due to their extreme biocompatibility. Unlike linear peptides, cyclic peptides have more structural rigidity and tolerance to enzymatic degradation and high environmental stability which make them even better candidates for the above-said applications. Herein, a new small cyclic dipeptide (CDP) cyclo-(Leu-S-Bzl-Cys) (P1) consisting of L-leucine and S-benzyl protected L-cysteine was reported which formed a hydrogel at physiological conditions (at 37°C and pH = 7.46). The hydrogel formed from the cyclic dipeptide P1 showed very good tolerance towards environmental parameters such as pH and temperature and was seen to be stable for more than a year without any deformation. The hydrogel was thermoreversible and stable in the pH range 6-12. Mechanical strength of P1 hydrogel was measured by rheology experiments. Atomic force microscopy (AFM) and field emission scanning electron microscopy (FE-SEM) images revealed that, in aqueous solvents, P1 self-assembled into a highly cross-linked nanofibrillar network which immobilized water molecules inside the cages and formed the hydrogel. The self-assembled cyclic dipeptide acquired the antiparallel β-sheet secondary structure, which was evident from CD and Fourier transform infrared (FT-IR) studies. The β-sheet arrangement and formation of amyloid fibrils were further established by ThT binding assay. Furthermore, P1 was able to form a hydrogel in the presence of the anticancer drug 5-fluorouracil (5FU), and sustainable release of the drug from the hydrogel was measured in vitro. The hydrogelator P1 showed almost no cytotoxicity towards the human colorectal cancer cell line HCT116 up to a considerably high concentration and showed potential application in sustainable drug delivery. The co-assembly of 5FU and P1 hydrogel exhibited much better anticancer activity towards the HCT116 cancer cell line than 5FU alone and decreased the IC50 dose of 5FU to a much lower value.

Marine Cyclic Dipeptide Cyclo (L-Leu-L-Pro) Protects Normal Breast Epithelial Cells from tBHP-induced Oxidative Damage by Targeting CD151

Background: Oxidative stress plays a key role in breast carcinogenesis. Cyclo (L-Leu-L-Pro) (CLP) is a homodetic cyclic dipeptide with 2,5-diketopiperazine scaffold isolated from marine actinobacteria. This study aimed to evaluate the protective activity of CLP and linear - (L-Leu-L-Pro) (LP) from tert-butyl hydroperoxide (tBHP)-induced damage using normal breast epithelial cell line model (MCF-12A). Methods: The cytoprotective activity was evaluated by detecting the changes in intracellular ROS, mitochondrial superoxide, hydroxyl radical, hydrogen peroxide, and lipid peroxidation detection assays as well as cytotoxic assays of MTT, LDH assays and phase contrast microscopy. Genoprotective activity was evaluated by (Apurinic/Apyrimidinic) AP site, alkaline Comet, and 8-hydroxy-2-deoxyguanosine assays. Results: The marine cyclic peptide, CLP, significantly protected MCF-12A cells by scavenging tBHP induced intracellular ROS such as super oxide, hydroxyl radicals and hydrogen peroxide, and by reducing the cytotoxicity and genotoxicity effect compared to LP. Moreover, the results showed that CD151 gene silencing by shRNA significantly reduced the overexpression of CD151, tBHP-induced ROS generation, cytotoxicity and genotoxicity in MCF-12A cells. The overexpression of CD151 caused increased levels of cytochrome P450, but was reduced following the application of CD151shRNA and CLP which led to elevated levels of intracellular ROS. Conclusion: In the present study we noticed that CD151 gene silencing by shRNA and treatment with CLP have similar effects on reducing the intracellular ROS. This study uncovers the protective activity of CLP against a CD151-mediated oxidative stress-induced cellular damage. Our observations suggest that the anti-stress and anti-inflammation properties of CLP might have implications in cancer and are worth testing in cancer cell lines and tumor cells.

Structure and collision-induced dissociation of the protonated cyclo His-Phe dipeptide: mechanistic studies and stereochemical effects

The role of stereochemical factors on the structure and the fragmentation paths of the protonated cyclic dipeptide cyclo histidine–phenylalanine is studied under ion traps conditions by combining tandem mass spectrometry, laser spectroscopy, quantum chemical calculations and chemical dynamics simulations. Vibrational spectroscopy obtained by Infrared Multiple Photon Dissociation (IRMPD) reveals a small difference between the two diastereomers, c- $$\hbox {LLH}^{+}$$ and c- $$\hbox {LDH}^{+}$$ , arising mainly from ancillary CH... $$\uppi $$ interactions. In contrast, there is a strong influence of the residues chirality on the collision-induced dissociation (CID) processes. Chemical dynamics simulations rationalize this effect and evidence that proton mobility takes place, allowing isomerization to intermediate cyclic structures that are different for c- $$\hbox {LLH}^{+}$$ and c- $$\hbox {LDH}^{+}$$ , resulting in different barriers to proton mobility. This effect is related to the protonation of the imidazole ring. It contrasts with the minute stereochemical effects observed for other cyclic dipeptides in which the proton is borne by an amide CO.

Biocontrol of the root-knot nematode Meloidogyne incognita by a nematicidal bacterium Pseudomonas simiae MB751 with cyclic dipeptide.

Root-knot nematodes (RKNs) are harmful plant-parasitic nematodes that cause serious damage to plant hosts. In the long-term practice of RKN management, bacterial nematicides have attracted increasing attention as an effective biocontrol means. Here we determined the active substances against Meloidogyne incognita from a nematicidal bacterium, developed a biocontrol agent (BCA) based on optimized culture processes. The effects of the BCA on RKN control and plant growth-promotion were evaluated in tomato pot trials. Pseudomonas simiae strain MB751 exhibiting significant nematicidal activity against M. incognita second-stage juveniles (J2) with approximately 80% mortality (with culture supernatant, 96% volume percentage) was isolated from a vineyard. A set of purification and identification experiments was performed to determine the main nematicidal component in MB751. A cyclic dipeptide Cyclo(L-Pro-L-Leu) was identified with a lethal concentration necessary to kill 50% of the population (LC50 ) of 65.3μg mL-1 against M. incognita J2. Following optimization trials on culture medium/fermentation conditions, such as the single factor test, Plackett-Burman test, steepest ascent, and response surface methodology experiments, the MB751 fermentation broth was then prepared as a BCA via a cold-air drying process. The BCA and was evaluated in tomato pot experiments for effectiveness in suppressing M. incognita. Significant effects on M. incognita suppression and plant-growth promotion as well as induced systemic resistance to M. incognita of tomato, were observed. The cyclic dipeptide-producing bacterium P. simiae MB751 exhibited high nematicidal activity and performance. Further development of this BCA should be pursued for the management of M. incognita in agriculture. © 2021 Society of Chemical Industry.

A cyclic dipeptide from the Chilean hazelnut cotyledons (Gevuina avellana Mol., Proteaceae)

The Chilean hazelnut (Gevuina avellana Mol., Proteaceae) is a southern South American nut consumed as a snack and included in different preparations of traditional Chilean cuisine. Recently we described the fatty acid profile, oxylipins, phenolic compounds, as well as the antioxidant capacity. The main compounds of the phenolic-enriched extract were only tentatively identified by spectrometric means. In the present work, we describe the isolation and full characterization of a cyclic dipeptide cyclo(Arg-Trp) and other compounds from the phenolic enriched extracts of the G. avellana cotyledons. Compounds were isolated by means of counter-current chromatography and structures were established by spectroscopic and spectrometric methods. This is the first report on small peptides in G. avellana and adds evidence on the possible beneficial effects of this nut in human health.

Reactive oxygen species generation and mitochondrial dysfunction for the initiation of apoptotic cell death in human hepatocellular carcinoma HepG2 cells by a cyclic dipeptide Cyclo(-Pro-Tyr).

Cyclic dipeptides are increasingly gaining importance as considering its significant biological and pharmacological activities. This study was aimed to investigate the anticancer activity of a dipeptide Cyclo(-Pro-Tyr) (DP) identified from marine sponge Callyspongia fistularis symbiont Bacillus pumilus AMK1 and the underlying apoptotic mechanisms in the liver cancer HepG2 cell lines. MTT assay was done to demonstrate the cytotoxic effect of DP in HepG2 cells and mouse Fibroblast McCoy cells. Initially, apoptosis inducing activity of DP was identified using propidium iodide (PI) and acridine orange/ethidium bromide (AO/EB) dual staining, then it was confirmed by DNA fragmentation assay and western blotting analysis of apoptosis related markers Bax, Bcl-2, cytochrome c, caspase-3 and cleaved poly (ADP-ribose) polymerase (PARP). Rhodamine 123 staining was performed to observe DP effects on the mitochondrial membrane potential (MMP) and DCFH-DA (Dichloro-dihydro-fluorescein diacetate) staining was done to measure the intracellular reactive oxygen species (ROS) levels. The MTT results revealed that DP initiated dose-dependent cytotoxicity in HepG2 cells, but no significant toxicity in mouse Fibroblast McCoy cells treated with DP at the specified concentrations. DP induced apoptosis, which is confirmed by the appearance of apoptotic bodies with PI and AO/EB dual staining, and DNA fragmentation. DP significantly elevated the Bax/Bcl-2 ratio, disrupted the mitochondrial membrane potential (MMP), enhanced cytochrome c release from mitochondria, increased caspase-3 activation, the cleavage of PARP and increased intracellular reactive oxygen species (ROS) levels. Besides this, DP successfully inhibited the phosphorylation of PI3K, AKT and increased PTEN expression. These results suggested DP might have anti-cancer effect by initiating apoptosis through mitochondrial dysfunction and downregulatingPI3K/Akt signaling pathway in HepG2 cells with no toxicity effect on normal fibroblast cells. Therefore, DP may be developed as a potential alternative therapeutic agent for treating hepatocellular carcinoma.

Screening of Various Metabolites in Six Barley Varieties Grown under Natural Climatic Conditions (2016-2018).

Climatic changes influence considerably the distribution and occurrence of different secondary metabolites in cereals. The aim of this investigation was to assess the changes in metabolite prevalence observed in six different winter barley varieties over a statistically significant period of three years by linking agro-climatic conditions with metabolite concentrations in chosen samples. The results showed that temperatures and precipitation levels varied during the observed timeframe and that the multi-toxin concentrations followed the trend of changing climatic conditions depending on the variety. All quantified (fungal) metabolites showed significant variations throughout the years and, for some (tryptophol and the cyclic dipeptides cyclo(L-Pro-L-Tyr) and cyclo(L-Pro-L-Val)), an unexpected, but clear connection can be made with temperature changes and precipitation levels during the growing season.

High-tolerance crystalline hydrogels formed from self-assembling cyclic dipeptide.

Peptide-based supramolecular hydrogels, as a new type of biological nanoarchitectonic structure, hold great promise for a wide range of biomedical and nanotechnological applications, such as tissue engineering, drug delivery, and electronic and photonic energy storage. In this work, a cyclic dipeptide (CDP) cyclo-(Trp-Tyr) (C-WY), which has exceptional structural rigidity and high stability, is selected as a hydrogelator for the formation of supramolecular hydrogels. The unique hydrogen bonding in C-WY endows a high propensity for self-assembly and the resulting hydrogels are revealed to be crystalline. The crystalline hydrogels possess excellent mechanical capacity and superior tolerance to various harsh conditions, including in the presence of charged biopolymers, extreme acid/base environments, and changing thermal conditions. Such high tolerance enables the crystalline hydrogels to be applied in the complex and harsh environments of electrochemistry. In addition, this study demonstrates that the self-assembly of cyclic dipeptides results in highly robust hydrogels which can be applied for electrochemical applications such as electrochemical supercapacitors.

Brevibacillus laterosporus isolated from the digestive tract of honeybees has high antimicrobial activity and promotes growth and productivity of honeybee's colonies.

The development of novel antimicrobial drugs, as well as the discovery of novel compounds able to promote honeybee's growth, represents major challenges for modern entomology. The main aim of this study was to investigate whether Brevibacillus laterosporus isolated from the digestive tract of Saudi honeybees, Apis mellifera, was able to stimulate colony strength parameters of honeybees and to evaluate its ability to produce antimicrobial agents. Honeybees were collected in Dirab, Riyadh Region, Saudi Arabia, and microorganisms were isolated and identified by 16S ribosomal RNA analysis. Microscopic identification of the microorganism in its native state was facilitated by atomic force microscopy at high-resolution imaging. Active biological compounds were produced by submerged fermentation with B. laterosporus. The fermented broth was subjected to extraction and purification, and then semi-pure compounds were analyzed by gas chromatography-mass spectrometry. The effectiveness of the crude extract and semi-pure compounds as antimicrobial agents was evaluated by susceptibility assays. More than 22% of the microorganisms isolated from the digestive tract of healthy honeybees have been identified as B. laterosporus, this kind of species has a unique shape and morphological structure. The cyclic dipeptide cyclo(Leu-Pro) produced by B. laterosporus showed biological activity against several pathogenic microorganisms. Furthermore, the total counts of workers, closed brood, and open brood, as well as the production of bee pollen and honey, were better in honeybees treated with a B. laterosporus suspension. The data indicated that the B. laterosporus strain isolated from a healthy honeybee might be a novel probiotic and a producer of important biological compounds.

Expanding the chemical space for natural products by Aspergillus-Streptomyces co-cultivation and biotransformation.

Actinomycetes and filamentous fungi produce a wide range of bioactive compounds, with applications as antimicrobials, anticancer agents or agrochemicals. Their genomes contain a far larger number of gene clusters for natural products than originally anticipated, and novel approaches are required to exploit this potential reservoir of new drugs. Here, we show that co-cultivation of the filamentous model microbes Streptomyces coelicolor and Aspergillus niger has a major impact on their secondary metabolism. NMR-based metabolomics combined with multivariate data analysis revealed several compounds that correlated specifically to co-cultures, including the cyclic dipeptide cyclo(Phe-Phe) and 2-hydroxyphenylacetic acid, both of which were produced by A. niger in response to S. coelicolor. Furthermore, biotransformation studies with o-coumaric acid and caffeic acid resulted in the production of the novel compounds (E)-2-(3-hydroxyprop-1-en-1-yl)-phenol and (2E,4E)-3-(2-carboxy-1-hydroxyethyl)-2,4-hexadienedioxic acid, respectively. This highlights the utility of microbial co-cultivation combined with NMR-based metabolomics as an efficient pipeline for the discovery of novel natural products.

Quantification of cyclic dipeptides from cultures of Lactobacillus brevis R2Δ by HRGC/MS using stable isotope dilution assay

Lactic acid bacteria (LAB) play an important role as natural preservatives in fermented food and beverage systems, reducing the application of chemical additives. Thus, investigating their antifungal compounds, such as cyclic dipeptides, has gained prominence. Previous research has primarily focussed on isolation of these compounds. However, their precise quantification will provide further information regarding their antifungal performance in a complex system. To address this, deuterated labelled standards of the cyclic dipeptides cyclo(Leu-Pro), cyclo(Pro-Pro), cyclo(Met-Pro) and cyclo(Phe-Pro) were synthesized, and stable isotope dilution assays were developed, enabling an accurate quantification of cyclo(Leu-Pro), cyclo(Pro-Pro), cyclo(Met-Pro) and cyclo(Phe-Pro) in MRS-broth and wort. Quantitative results showed that, in the Lactobacillus brevis R2Δ fermented MRS-broth, the concentrations of cyclo(Leu-Pro), cyclo(Pro-Pro) and cyclo(Phe-Pro) were significantly higher (P < 0.05), than in wort for cyclo (Leu-Pro) when compared with their controls. This indicates that the formation of these three cyclic dipeptides is related to L. brevis R2Δ metabolism. Furthermore, this represents the first report of cyclic dipeptides quantification using stable isotope dilution assays in LAB cultures both in vitro and in a food system.

Transformation of Gold(I)–cyclo[–Met–Met–] Complex Supramolecular Fibers into Aligned Gold Nanoparticles

Abstract A cyclic dipeptide cyclo[–l-methionyl–l-methionyl–] and gold(I) cation were found to form a complex in a 1:2 ratio. Pouring a DMF solution of the complex into ethyl acetate afforded fibers with lengths of more than 20 µm and 100–400 nm in width, which consisted of fibrous nanostructures 50–100 nm in diameter. Treatment of this supramolecular fiber with catechol gave linearly aligned Au nanoparticles on the original fibers.

Determination of Absolute Configuration and Conformation of a Cyclic Dipeptide by NMR and Chiral Spectroscopic Methods

Increasing precision of contemporary computational methods makes spectroscopies such as vibrational (VCD) and electronic (ECD) circular dichroism attractive for determination of absolute configurations (AC) of organic compounds. This is, however, difficult for polar, flexible molecules with multiple chiral centers. Typically, a combination of several methods provides the best picture of molecular behavior. As a test case, all possible stereoisomers with known AC (RS, SR, SS, and RR) of the cyclic dipeptide cyclo(Arg-Trp) (CAT) were synthesized, and the performances of the ECD, infrared (IR), VCD, Raman, Raman optical activity (ROA), and nuclear magnetic resonance (NMR) techniques for AC determination were investigated. The spectra were interpreted with the aid of density functional theory (DFT) calculations. Folded geometries stabilized by van der Waals and electrostatic interactions between the diketopiperazine (DKP) ring and the indole group are predicted to be preferred for CAT, with more pronounced folding due to Arg-Trp stacking in the case of SS/RR-CAT. The RS/SR isomers prefer a twist-boat puckering of the DKP ring, which is relatively independent of the orientation of the side chains. Calculated conformer-averaged VCD and ECD spectra explain most of the experimentally observed bands and allow for AC determination of the tryptophan side-chain, whereas the stereochemical configuration of the arginine side-chain is visible only in VCD. NMR studies provide characteristic long-range (2)J(C,H) and (3)J(C,H) coupling constants, and nuclear Overhauser effect (NOE) correlations, which in combination with either ECD or VCD also allow for complete AC determination of CAT.

Isolation of proline-based cyclic dipeptides from Bacillus sp. N strain associated with rhabitid entomopathogenic nematode and its antimicrobial properties

Entomopathogenic nematodes (EPN) are well-known as biological control agents and are found to have associated bacteria which can produce a wide range of bioactive secondary metabolites. We report herewith isolation of six proline containing cyclic dipeptides cyclo(D-Pro-L-Leu), cyclo(L-Pro-L-Met), cyclo(D-Pro-L-Phe), cyclo(L-Pro-L-Phe), cyclo(L-Pro-L-Tyr) and cyclo(L-Pro-D-Tyr) from ethyl acetate extract of the Luria Broth (LB) cell free culture filtrate of Bacillus sp. strain N associated with a new EPN Rhabditis sp. from sweet potato weevil grubs collected from Central Tuber Crops Research Institute farm. Antimicrobial studies of these 2,5-diketopiperazines (DKPs) against both medicinally and agriculturally important bacterium and fungi showed potent inhibitory values in the range of μg/mL. Cyclic dipeptides showed significantly higher activity than the commercial fungicide bavistin against agriculturally important fungi, viz., Fusarium oxysporum, Rhizoctonia solani, and Pencillium expansum. The highest activity of 2 μg/mL by cyclo(L-Pro-L-Phe) was recorded against P. expansum, a plant pathogen responsible for causing post harvest decay of stored apples and oranges. To our knowledge, this is the first report on the isolation of these DKPs from Rhabditis EPN bacterial strain Bacillus sp.

Lactobacillus reuteri -produced cyclic dipeptides quench agr -mediated expression of toxic shock syndrome toxin-1 in staphylococci

The production of the staphylococcal exotoxin toxic shock syndrome toxin-1 (TSST-1) by Staphylococcus aureus has been associated with essentially all cases of menstruation-associated toxic shock syndrome (TSS). In this work, we show that the human vaginal isolate Lactobacillus reuteri RC-14 produces small signaling molecules that are able to interfere with the staphylococcal quorum-sensing system agr, a key regulator of virulence genes, and repress the expression of TSST-1 in S. aureus MN8, a prototype of menstrual TSS S. aureus strains. Quantitative real-time PCR data showed that transcription from the Ptst promoter, as well as the P2 and P3 promoters of the agr system from all four agr subgroups of S. aureus, was strongly inhibited in response to growth with L. reuteri RC-14 cultural supernatant. Alterations in the transcriptional levels of two other virulence-associated regulators sarA and saeRS were also observed, indicating a potential overall influence of L. reuteri RC-14 signals on the production of virulence factors in S. aureus. S. aureus promoter-lux reporter strains were used to screen biochemically fractionated L. reuteri RC-14 supernatant, and the cyclic dipeptides cyclo(L-Phe-L-Pro) and cyclo(L-Tyr-L-Pro) were identified as the signaling molecules. The results from this work contribute to a better understanding of interspecies cell-to-cell communication between Lactobacillus and Staphylococcus, and provide a unique mechanism by which endogenous or probiotic strains may attenuate virulence factor production by bacterial pathogens.