The Experts below are selected from a list of 360 Experts worldwide ranked by ideXlab platform
Dejing Shang - One of the best experts on this subject based on the ideXlab platform.
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antimicrobial peptide Temporin 1cea isolated from frog skin secretions inhibits the proinflammatory response in lipopolysaccharide stimulated raw264 7 murine macrophages through the myd88 dependent signaling pathway
Molecular Immunology, 2021Co-Authors: Juan Zhang, Yue Sun, Yao Kang, Dejing ShangAbstract:Temporin-1CEa, which is isolated from the skin secretions of the Chinese brown frog Rana chensinensis, exhibits broad-spectrum antimicrobial activity against gram-positive and gram-negative bacteria and antitumor activity. LK2(6) and LK2(6)A(L) are the analogs of Temporin-1CEa obtained by replacing amino acids and displayed an improved anticancer activity. In the present study, the anti-inflammatory activity and mechanism of action of Temporin-1CEa and its analogs LK2(6) and LK2(6)A(L) in lipopolysaccharide (LPS)-stimulated RAW264.7 murine macrophages were investigated. The results showed that Temporin-1CEa and its analogs decreased the production of the cytokines tumor necrosis factor-α and interleukin-6 by inhibiting the protein expression of nuclear factor-κB and mitogen-activated protein kinase and the MyD88-dependent signaling pathway. Isothermal titration calorimetry studies revealed that Temporin-1CEa, LK2(6) and LK2(6)A(L) exhibited binding affinities to LPS, an important inflammatory inducer, with Kd values of 0.1, 0.03 and 0.06 μM, respectively. Circular dichroism and zeta potential experiments showed that Temporin-1CEa and its analogs interacted with LPS by electrostatic binding between the positively charged peptides and negatively charged LPS, resulting in the neutralization of LPS toxicity.
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melanoma cell surface expressed phosphatidylserine as a therapeutic target for cationic anticancer peptide Temporin 1cea
Journal of Drug Targeting, 2016Co-Authors: Che Wang, Yang Zhou, Yinwang Chen, Liang Zhang, Xiange Gong, Dejing ShangAbstract:We have previously reported that Temporin-1CEa, a cationic antimicrobial peptide, exerts preferential cytotoxicity toward cancer cells. However, the exact molecular mechanism for this cancer-selectivity is still largely unknown. Here, we found that the negatively charged phosphatidylserine (PS) expressed on cancer cell surface serves as a target for Temporin-1CEa. Our results indicate that human A375 melanoma cells express 50-fold more PS than non-cancerous HaCaT cells. The expression of cell surface PS in various cancer cell lines closely correlated with their ability to be recognized, bound and killed by Temporin-1CEa. Additionally, the cytotoxicity of Temporin-1CEa against A375 cells can be ameliorated by annexin V, which binds to cell surface PS with high affinity. Moreover, the data of isothermal titration calorimetry assay further confirmed a direct binding of Temporin-1CEa to PS, at a ratio of 1:5 (Temporin-1CEa:PS). Interestingly, the circular dichroism spectra analysis using artificial biomembrane revealed that PS not only provides electrostatic attractive sites for Temporin-1CEa but also confers the membrane-bound Temporin-1CEa to form α-helical structure, therefore, enhances the affinity and membrane disrupting ability of Temporin-1CEa. In summary, these findings suggested that the melanoma cells expressed PS may serve as a promising target for Temporin-1CEa or other cationic anticancer peptides.
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anticancer mechanisms of Temporin 1cea an amphipathic α helical antimicrobial peptide in bcap 37 human breast cancer cells
Life Sciences, 2013Co-Authors: Che Wang, He Wang, Yang Zhou, Lili Tian, Dejing ShangAbstract:Abstract Aims Temporin-1CEa, a 17-residue antimicrobial peptide, is known to exert broad-spectrum anticancer activity that acts preferentially on cancer cells instead of normal cells. However, the mechanism of cancer cell death induced by Temporin-1CEa is weakly understood. Main methods Here, we investigated the cytotoxic and membrane-disrupting effects of Temporin-1CEa on human breast cancer cell line Bcap-37, using MTT assay, electronic microscope observation, fluorescence imaging and flow cytometry analysis. Key findings The MTT assay indicated that one-hour Temporin-1CEa treatment led to rapid cell death in either caspase-dependent or -independent manner. The electronic microscope observation suggested that Temporin-1CEa exposure resulted in profound morphological changes in Bcap-37 cells. The fluorescence imaging and flow cytometry analysis demonstrated that Temporin-1CEa exhibited membrane-disrupting property characterized by induction of cell-surface phosphatidylserine exposure, elevation of plasma membrane permeability, and rapid transmembrane potential depolarization. Moreover, Temporin-1CEa might also induce rapid cell death through mitochondria-involved mechanisms, including rapid intracellular Ca2 + leakage, collapse of mitochondrial membrane potential (Δφm) and over-generation of reactive oxygen species (ROS). Significance In summary, the present study indicates that Temporin-1CEa triggers a rapid cytotoxicity in Bcap-37 cells through membrane-destruction and intracellular mechanisms involving mitochondria. These intracellular mechanisms and direct membrane-destruction effect were evaluated helping to understand the detail action of antimicrobial peptides in mammalian cancer cells.
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design of potent non toxic anticancer peptides based on the structure of the antimicrobial peptide Temporin 1cea
Archives of Pharmacal Research, 2013Co-Authors: Qingzhu Yang, Che Wang, He Wang, Lei Lang, Yang Zhou, Dejing ShangAbstract:Recent advances in the search for novel anticancer agents have indicated that the positively charged antimicrobial peptides have emerged as promising agents offering several advantages over the conventional anticancer drugs. As a naturally occurring, cationic, α-helical antimicrobial peptide, temproin-1CEa has been proved to exhibit a potent anticancer effect and a moderate hemolytic activity. In order to reduce the hemolytic activity of Temporin-1CEa and improve its anticancer potency towards a range of human breast cancer cells, in the present study, six analogs of Temporin-1CEa were rationally designed and synthesized. The amphipathicity levels and α-helical structural patterns of peptides were reserved, while their cationic property and hydrophobicity were changed. The results of MTT and hemolysis assay indicated that the analog peptides displayed an improved anticancer activity and showed an overall optimized therapeutic index. The hydrophobicity of peptides was positively correlated with their hemolytic and antitumor activities. Moreover, the data suggest a strategy of increasing the cationicity while maintaining the moderate hydrophobicity of naturally occurring amphipathic α-helical peptides to generate analogs with improved cytotoxicity against tumor cells but decreased activity against non-neoplastic cells such as human erythrocytes. This work highlights the potential for rational design and synthesis of improved antimicrobial peptides that have the capability to be used therapeutically for treatment of cancers.
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design of potent non toxic antimicrobial agents based upon the structure of the frog skin peptide Temporin 1ceb from chinese brown frog rana chensinensis
Chemical Biology & Drug Design, 2012Co-Authors: Dejing Shang, Xiaofan Li, Che WangAbstract:Temporin-1CEb shows antimicrobial activity against Gram-positive bacteria, but its therapeutic potential is limited by its haemolysis. In this study, eight Temporin-1CEb analogues with altered cationicities and hydrophobicities were synthesized. Increasing cationicity and amphipathicity by substituting neutral and non-polar amino acid residues on the hydrophilic face of the α-helix by five or six lysines increased antimicrobial potency approximately 10-fold to 40-fold, although when the number of positive charges was increased from +6 to +7, the antimicrobial potency was not additionally enhanced. The substitution of an l-lysine with a d-lysine, meanwhile maintaining the net charge and the mean hydrophobicity values, had only a minor effect on its antimicrobial activity, whereas significantly led a decrease in its haemolytic activity. Of all the peptides, l-K6 has the best potential as an antimicrobial agent because its antimicrobial activity against both Gram-positive and Gram-negative bacteria is substantial, and its haemolytic activity is negligible. l-K6 adopts an α-helix in 50% trifluoroethanol/water and 30 mm SDS solutions. l-K6 killed 99.9% of E. coli and S. aureus at 4× MIC in 60 min, and its postantibiotic effect was >5 h. l-K6 affects the integrity of E. coli and S. aureus plasma membranes by rapidly inducing membrane depolarization.
Maria Luisa Mangoni - One of the best experts on this subject based on the ideXlab platform.
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Temporin g an amphibian antimicrobial peptide against influenza and parainfluenza respiratory viruses insights into biological activity and mechanism of action
The FASEB Journal, 2021Co-Authors: M De Angelis, Bruno Casciaro, Maria Luisa Mangoni, Ettore Novellino, Diego Brancaccio, A Genovese, Maria Elena Marcocci, Antonio Carotenuto, Anna Teresa Palamara, Lucia NencioniAbstract:Treatment of respiratory viral infections remains a global health concern, mainly due to the inefficacy of available drugs. Therefore, the discovery of novel antiviral compounds is needed; in this context, antimicrobial peptides (AMPs) like Temporins hold great promise. Here, we discovered that the harmless Temporin G (TG) significantly inhibited the early life-cycle phases of influenza virus. The in vitro hemagglutinating test revealed the existence of TG interaction with the viral hemagglutinin (HA) protein. Furthermore, the hemolysis inhibition assay and the molecular docking studies confirmed a TG/HA complex formation at the level of the conserved hydrophobic stem groove of HA. Remarkably, these findings highlight the ability of TG to block the conformational rearrangements of HA2 subunit, which are essential for the viral envelope fusion with intracellular endocytic vesicles, thereby neutralizing the virus entry into the host cell. In comparison, in the case of parainfluenza virus, which penetrates host cells upon a membrane-fusion process, addition of TG to infected cells provoked ~1.2 log reduction of viral titer released in the supernatant. Nevertheless, at the same condition, an immunofluorescent assay showed that the expression of viral hemagglutinin/neuraminidase protein was not significantly reduced. This suggested a peptide-mediated block of some late steps of viral replication and therefore the impairment of the extracellular release of viral particles. Overall, our results are the first demonstration of the ability of an AMP to interfere with the replication of respiratory viruses with a different mechanism of cell entry and will open a new avenue for the development of novel therapeutic approaches against a large variety of respiratory viruses, including the recent SARS-CoV2.
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assessment of the potential of Temporin peptides from the frog rana temporaria ranidae as anti diabetic agents
Journal of Peptide Science, 2018Co-Authors: Vishal Musale, Yasser Abdelwahab, Peter R Flatt, Bruno Casciaro, Maria Luisa Mangoni, Michael J ConlonAbstract:Temporin A (FLPLIGRVLSGIL-NH2 ), Temporin F (FLPLIGKVLSGIL-NH2 ), and Temporin G (FFPVIGRILNGIL-NH2 ), first identified in skin secretions of the frog Rana temporaria, produced concentration-dependent stimulation of insulin release from BRIN-BD11 rat clonal β-cells at concentrations ≥1 nM, without cytotoxicity at concentrations up to 3 μM. Temporin A was the most effective. The mechanism of insulinotropic action did not involve an increase in intracellular Ca2+ concentrations. Temporins B, C, E, H, and K were either inactive or only weakly active. Temporins A, F, and G also produced a concentration-dependent stimulation of insulin release from 1.1B4 human-derived pancreatic β-cells, with Temporin G being the most potent and effective, and from isolated mouse islets. The data indicate that cationicity, hydrophobicity, and the angle subtended by the charged residues in the Temporin molecule are important determinants for in vitro insulinotropic activity. Temporin A and F (1 μM), but not Temporin G, protected BRIN-BD11 cells against cytokine-induced apoptosis (P < 0.001) and augmented (P < 0.001) proliferation of the cells to a similar extent as glucagon-like peptide-1. Intraperitoneal injection of Temporin G (75 nmol/kg body weight) together with a glucose load (18 mmol/kg body weight) in C57BL6 mice improved glucose tolerance with a concomitant increase in insulin secretion whereas Temporin A and F administration was without significant effect on plasma glucose levels. The study suggests that combination therapy involving agents developed from the Temporin A and G sequences may find application in Type 2 diabetes treatment.
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nmr structure of Temporin 1 ta in lipopolysaccharide micelles mechanistic insight into inactivation by outer membrane
PLOS ONE, 2013Co-Authors: Rathi Saravanan, Harini Mohanram, Maria Luisa Mangoni, Anirban Bhunia, Mangesh Joshi, Surajit BhattacharjyaAbstract:Background: Antimicrobial peptides (AMPs) play important roles in the innate defense mechanism. The broad spectrum of activity of AMPs requires an efficient permeabilization of the bacterial outer and inner membranes. The outer leaflet of the outer membrane of Gram negative bacteria is made of a specialized lipid called lipopolysaccharide (LPS). The LPS layer is an efficient permeability barrier against anti-bacterial agents including AMPs. As a mode of protection, LPS can induce self associations of AMPs rendering them inactive. Temporins are a group of short-sized AMPs isolated from frog skin, and many of them are inactive against Gram negative bacteria as a result of their self-association in the LPS-outer membrane. Principal Findings: Using NMR spectroscopy, we have determined atomic resolution structure and characterized localization of Temporin-1Ta or TA (FLPLIGRVLSGIL-amide) in LPS micelles. In LPS micelles, TA adopts helical conformation for residues L4-I12, while residues F1-L3 are found to be in extended conformations. The aromatic sidechain of residue F1 is involved in extensive packing interactions with the sidechains of residues P3, L4 and I5. Interestingly, a number of longrange NOE contacts have been detected between the N-terminal residues F1, P3 with the C-terminal residues S10, I12, L13 of TA in LPS micelles. Saturation transfer difference (STD) NMR studies demonstrate close proximity of residues including F1, L2, P3, R7, S10 and L13 with the LPS micelles. Notably, the LPS bound structure of TA shows differences with the structures of TA determined in DPC and SDS detergent micelles. Significance: We propose that TA, in LPS lipids, forms helical oligomeric structures employing N- and C-termini residues. Such oligomeric structures may not be translocated across the outer membrane; resulting in the inactivation of the AMP. Importantly, the results of our studies will be useful for the development of antimicrobial agents with a broader spectrum of activity.
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nmr structures and interactions of Temporin 1tl and Temporin 1tb with lipopolysaccharide micelles mechanistic insights into outer membrane permeabilization and synergistic activity
Journal of Biological Chemistry, 2011Co-Authors: Anirban Bhunia, Harini Mohanram, Maria Luisa Mangoni, Rathi Saravanan, Surajit BhattacharjyaAbstract:Temporins are a group of closely related short antimicrobial peptides from frog skin. Lipopolysaccharide (LPS), the major constituent of the outer membrane of gram-negative bacteria, plays important roles in the activity of Temporins. Earlier studies have found that LPS induces oligomerization of Temporin-1Tb (TB) thus preventing its translocation across the outer membrane and, as a result, reduces its activity on gram-negative bacteria. On the other hand, Temporin-1Tl (TL) exhibits higher activity, presumably because of lack of such oligomerization. A synergistic mechanism was proposed, involving TL and TB in overcoming the LPS-mediated barrier. Here, to gain insights into interactions of TL and TB within LPS, we investigated the structures and interactions of TL, TB, and TL+TB in LPS micelles, using NMR and fluorescence spectroscopy. In the context of LPS, TL assumes a novel antiparallel dimeric helical structure sustained by intimate packing between aromatic-aromatic and aromatic-aliphatic residues. By contrast, independent TB has populations of helical and aggregated conformations in LPS. The LPS-induced aggregated states of TB are largely destabilized in the presence of TL. Saturation transfer difference NMR studies have delineated residues of TL and TB in close contact with LPS and enhanced interactions of these two peptides with LPS, when combined together. Fluorescence resonance energy transfer and (31)P NMR have pointed out the proximity of TL and TB in LPS and conformational changes of LPS, respectively. Importantly, these results provide the first structural insights into the mode of action and synergism of antimicrobial peptides at the level of the LPS-outer membrane.
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alanine scanning analysis and structure function relationships of the frog skin antimicrobial peptide Temporin 1ta
Journal of Peptide Science, 2011Co-Authors: Paolo Grieco, Pietro Campiglia, Donatella Barra, Vincenzo Luca, Luigia Auriemma, Alfonso Carotenuto, Maria Rosaria Saviello, Ettore Novellino, Maria Luisa MangoniAbstract:The increasing resistance of bacteria and fungi to the available antibiotic/antimycotic drugs urges for a search for new anti-infective compounds with new modes of action. In line of this, natural CAMPs represent promising and attractive candidates. Special attention has been devoted to frog-skin Temporins, because of their short size (10-14 residues long) and their unique features. In particular, Temporin-1Ta has the following properties: (i) it is mainly active on Gram-positive bacteria; (ii) it can synergize, when combined with Temporin-1Tl, in inhibiting both gram-negative bacterial growth and the toxic effect of LPS; (iii) it preserves biological activity in the presence of serum; and (iv) it is practically not hemolytic. Rational design of CAMPs represents a straightforward approach to obtain a peptide with a better therapeutic index. Here, we used alanine scanning analogs to elucidate the contribution of the side chains of each amino acid residue to the peptide's antimicrobial and hemolytic activity. Beside providing insight into the biophysical attributes and the critical positions within the peptide sequence, which govern the antimicrobial/hemolytic activity of this Temporin isoform, our studies assist in optimizing the design of Temporin-based lead structures for the production of new anti-infective agents.
Che Wang - One of the best experts on this subject based on the ideXlab platform.
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melanoma cell surface expressed phosphatidylserine as a therapeutic target for cationic anticancer peptide Temporin 1cea
Journal of Drug Targeting, 2016Co-Authors: Che Wang, Yang Zhou, Yinwang Chen, Liang Zhang, Xiange Gong, Dejing ShangAbstract:We have previously reported that Temporin-1CEa, a cationic antimicrobial peptide, exerts preferential cytotoxicity toward cancer cells. However, the exact molecular mechanism for this cancer-selectivity is still largely unknown. Here, we found that the negatively charged phosphatidylserine (PS) expressed on cancer cell surface serves as a target for Temporin-1CEa. Our results indicate that human A375 melanoma cells express 50-fold more PS than non-cancerous HaCaT cells. The expression of cell surface PS in various cancer cell lines closely correlated with their ability to be recognized, bound and killed by Temporin-1CEa. Additionally, the cytotoxicity of Temporin-1CEa against A375 cells can be ameliorated by annexin V, which binds to cell surface PS with high affinity. Moreover, the data of isothermal titration calorimetry assay further confirmed a direct binding of Temporin-1CEa to PS, at a ratio of 1:5 (Temporin-1CEa:PS). Interestingly, the circular dichroism spectra analysis using artificial biomembrane revealed that PS not only provides electrostatic attractive sites for Temporin-1CEa but also confers the membrane-bound Temporin-1CEa to form α-helical structure, therefore, enhances the affinity and membrane disrupting ability of Temporin-1CEa. In summary, these findings suggested that the melanoma cells expressed PS may serve as a promising target for Temporin-1CEa or other cationic anticancer peptides.
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anticancer mechanisms of Temporin 1cea an amphipathic α helical antimicrobial peptide in bcap 37 human breast cancer cells
Life Sciences, 2013Co-Authors: Che Wang, He Wang, Yang Zhou, Lili Tian, Dejing ShangAbstract:Abstract Aims Temporin-1CEa, a 17-residue antimicrobial peptide, is known to exert broad-spectrum anticancer activity that acts preferentially on cancer cells instead of normal cells. However, the mechanism of cancer cell death induced by Temporin-1CEa is weakly understood. Main methods Here, we investigated the cytotoxic and membrane-disrupting effects of Temporin-1CEa on human breast cancer cell line Bcap-37, using MTT assay, electronic microscope observation, fluorescence imaging and flow cytometry analysis. Key findings The MTT assay indicated that one-hour Temporin-1CEa treatment led to rapid cell death in either caspase-dependent or -independent manner. The electronic microscope observation suggested that Temporin-1CEa exposure resulted in profound morphological changes in Bcap-37 cells. The fluorescence imaging and flow cytometry analysis demonstrated that Temporin-1CEa exhibited membrane-disrupting property characterized by induction of cell-surface phosphatidylserine exposure, elevation of plasma membrane permeability, and rapid transmembrane potential depolarization. Moreover, Temporin-1CEa might also induce rapid cell death through mitochondria-involved mechanisms, including rapid intracellular Ca2 + leakage, collapse of mitochondrial membrane potential (Δφm) and over-generation of reactive oxygen species (ROS). Significance In summary, the present study indicates that Temporin-1CEa triggers a rapid cytotoxicity in Bcap-37 cells through membrane-destruction and intracellular mechanisms involving mitochondria. These intracellular mechanisms and direct membrane-destruction effect were evaluated helping to understand the detail action of antimicrobial peptides in mammalian cancer cells.
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purification molecular cloning and antimicrobial activity of peptides from the skin secretion of the black spotted frog rana nigromaculata
World Journal of Microbiology & Biotechnology, 2013Co-Authors: Yong Zhang, Che Wang, Zhenchun WangAbstract:Antimicrobial peptides from a wide range of amphibian species, especially frogs of the genus Rana, have been characterised and are potential therapeutic agents. Here we describe the isolation, purification, and structural and biological characterisation of three novel antimicrobial peptides from the skin secretions of the black spotted frog, Rana nigromaculata, from Northeastern China. The peptides were identified as belonging to two known families: the Temporin, which was first identified in R. nigromaculata from China, and the brevinin-2. Temporin-1RNa and Temporin-1RNb both containing three positive charges and have a high potency against microorganisms (MIC: 3.13–8.3 μM against Gram-positive bacteria, 12.5–25.0 μM against Gram-negative bacteria, and 6.25–12.5 μM against Candida albicans) and a high haemolytic activity against human erythrocytes (HC50: 100–150 μM). Brevinin-2RNa contains a single intra-disulphide bridge at the C-terminus that is active towards the tested Gram-positive bacteria but is not active against E. coli and P. aeruginosa. The cDNAs encoding three novel peptide precursors were also subsequently cloned from an R. nigromaculata skin cDNA library and sequenced. The precursors contain 58–72 amino acid residues, which include a conserved signal peptide, acidic propeptide, and the mature Temporin-1RNa, Temporin-1RNb and brevinin-2RNa. The CD spectra of Temporin-1RNa and Temporin-1RNb in water, 30 mM SDS and 50 % trifluoroethanol (TFE) indicated that both peptides adopted an aperiodic structure in water and an organised structure with an α-helical conformation in TFE and SDS solution. The conformational transition induced by TFE or SDS reflects the potential ability of Temporin-1RNa and Temporin-1RNb to interact with anionic membranes.
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design of potent non toxic anticancer peptides based on the structure of the antimicrobial peptide Temporin 1cea
Archives of Pharmacal Research, 2013Co-Authors: Qingzhu Yang, Che Wang, He Wang, Lei Lang, Yang Zhou, Dejing ShangAbstract:Recent advances in the search for novel anticancer agents have indicated that the positively charged antimicrobial peptides have emerged as promising agents offering several advantages over the conventional anticancer drugs. As a naturally occurring, cationic, α-helical antimicrobial peptide, temproin-1CEa has been proved to exhibit a potent anticancer effect and a moderate hemolytic activity. In order to reduce the hemolytic activity of Temporin-1CEa and improve its anticancer potency towards a range of human breast cancer cells, in the present study, six analogs of Temporin-1CEa were rationally designed and synthesized. The amphipathicity levels and α-helical structural patterns of peptides were reserved, while their cationic property and hydrophobicity were changed. The results of MTT and hemolysis assay indicated that the analog peptides displayed an improved anticancer activity and showed an overall optimized therapeutic index. The hydrophobicity of peptides was positively correlated with their hemolytic and antitumor activities. Moreover, the data suggest a strategy of increasing the cationicity while maintaining the moderate hydrophobicity of naturally occurring amphipathic α-helical peptides to generate analogs with improved cytotoxicity against tumor cells but decreased activity against non-neoplastic cells such as human erythrocytes. This work highlights the potential for rational design and synthesis of improved antimicrobial peptides that have the capability to be used therapeutically for treatment of cancers.
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design of potent non toxic antimicrobial agents based upon the structure of the frog skin peptide Temporin 1ceb from chinese brown frog rana chensinensis
Chemical Biology & Drug Design, 2012Co-Authors: Dejing Shang, Xiaofan Li, Che WangAbstract:Temporin-1CEb shows antimicrobial activity against Gram-positive bacteria, but its therapeutic potential is limited by its haemolysis. In this study, eight Temporin-1CEb analogues with altered cationicities and hydrophobicities were synthesized. Increasing cationicity and amphipathicity by substituting neutral and non-polar amino acid residues on the hydrophilic face of the α-helix by five or six lysines increased antimicrobial potency approximately 10-fold to 40-fold, although when the number of positive charges was increased from +6 to +7, the antimicrobial potency was not additionally enhanced. The substitution of an l-lysine with a d-lysine, meanwhile maintaining the net charge and the mean hydrophobicity values, had only a minor effect on its antimicrobial activity, whereas significantly led a decrease in its haemolytic activity. Of all the peptides, l-K6 has the best potential as an antimicrobial agent because its antimicrobial activity against both Gram-positive and Gram-negative bacteria is substantial, and its haemolytic activity is negligible. l-K6 adopts an α-helix in 50% trifluoroethanol/water and 30 mm SDS solutions. l-K6 killed 99.9% of E. coli and S. aureus at 4× MIC in 60 min, and its postantibiotic effect was >5 h. l-K6 affects the integrity of E. coli and S. aureus plasma membranes by rapidly inducing membrane depolarization.
Tianbao Chen - One of the best experts on this subject based on the ideXlab platform.
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aggregation and its influence on the bioactivities of a novel antimicrobial peptide Temporin pf and its analogues
International Journal of Molecular Sciences, 2021Co-Authors: Mei Zhou, Tianbao Chen, Lei Wang, Yu Zai, Xiaoling Chen, Shirley W I Siu, Hang Fai KwokAbstract:Temporin is an antimicrobial peptide (AMP) family discovered in the skin secretion of ranid frog that has become a promising alternative for conventional antibiotic therapy. Herein, a novel Temporin peptide, Temporin-PF (TPF), was successfully identified from Pelophylax fukienensis. It exhibited potent activity against Gram-positive bacteria, but no effect on Gram-negative bacteria. Additionally, TPF exhibited aggregation effects in different solutions. Three analogs were further designed to study the relationship between the aggregation patterns and bioactivities, and the MD simulation was performed for revealing the pattern of the peptide assembly. As the results showed, all peptides were able to aggregate in the standard culture media and salt solutions, especially CaCl2 and MgCl2 buffers, where the aggregation was affected by the concentration of the salts. MD simulation reported that all peptides were able to form oligomers. The parent peptide assembly depended on the hydrophobic interaction via the residues in the middle domain of the sequence. However, the substitution of Trp/D-Trp resulted in an enhanced inter-peptide interaction in the zipper-like domain and eliminated overall biological activities. Our study suggested that introducing aromaticity at the zipper-like domain for Temporin may not improve the bioactivities, which might be related to the formation of aggregates via the inter-peptide contacts at the zipper-like motif domain, and it could reduce the binding affinity to the lipid membrane of microorganisms.
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identification and target modifications of Temporin pe a novel antimicrobial peptide in the defensive skin secretions of the edible frog pelophylax kl esculentus
Biochemical and Biophysical Research Communications, 2018Co-Authors: Mengru Sang, James F. Burrows, Qinan Wu, Xinping Xi, Mei Zhou, Lei Wang, Tianbao ChenAbstract:Abstract A potent natural antimicrobial peptide named Temporin-PE was identified and encoded from the skin secretions of Pelophylax kl. esculentus via “shotgun” cloning and LC-MS/MS fragmentation analysis. Target-modifications were carried out to further enhance the antimicrobial and anti-proliferative bioactivities, whilst decreasing the hemolytic effect. A range of bioassays demonstrated that replacing a proline with a tyrosine residue resulted in a loss of the bioactivity against Gram-negative bacteria, but dramatically improved the hemolytic and anti-proliferative activity, indicating the FLP- motif influences the hemolytic activity of Temporins. Moreover, the coupling of TAT to the peptide dramatically improved its antimicrobial activity, indicating coupling TAT to these peptides could be considered as a potential tool to improve their antimicrobial activity. Overall, we have shown that targeted modifications of this natural antimicrobial peptide can adjust its bioactivities to help its development as an antibiotic or anti-proliferative agent.
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identification and bioactivity evaluation of two novel Temporins from the skin secretion of the european edible frog pelophylax kl esculentus
Biochemical and Biophysical Research Communications, 2016Co-Authors: Xiaole Chen, Mei Zhou, Tianbao Chen, Lei Wang, He Wang, Mu Yang, C. ShawAbstract:The amphibian Temporins, amongst the smallest antimicrobial peptides (AMPs), are α-helical, amphipathic, hydrophobic and cationic and are active mainly against Gram-positive bacteria but inactive or weakly active against Gram-negative bacteria. Here, we report two novel members of the Temporin family, named Temporin-1Ee (FLPVIAGVLSKLFamide) and Temporin-1Re (FLPGLLAGLLamide), whose biosynthetic precursor structures were deduced from clones obtained from skin secretion-derived cDNA libraries of the European edible frog, Pelophylax kl. esculentus, by 'shotgun' cloning. Deduction of the molecular masses of each mature processed peptide from respective cloned cDNAs was used to locate respective molecules in reverse-phase HPLC fractions of secretion. Temporin-1Ee (MIC = 10 μM) and Temporin-1Re (MIC = 60 μM) were both found to be active against Gram-positive Staphylococcus aureus, but retaining a weak haemolytic activity. To our knowledge, Single-site substitutions can dramatically change the spectrum of activity of a given Temporin. Compared with Temporine-1Ec, just one chemically-conservative substitution (Val8 instead of Leu8), Temporin-1Ee bearing a net charge of +2 displays broad-spectrum activity with particularly high potency on the clinically relevant Gram-negative strains, Escherichia coli (MIC = 40 μM). These factors bode well for translating Temporins to be potential drug candidates for the design of new and valuable anti-infective agents.
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identification and bioactivity evaluation of two novel Temporins from the skin secretion of the european edible frog pelophylax kl esculentus
Biochemical and Biophysical Research Communications, 2016Co-Authors: Xiaole Chen, Mei Zhou, Tianbao Chen, Lei Wang, He Wang, Mu Yang, C. ShawAbstract:The amphibian Temporins, amongst the smallest antimicrobial peptides (AMPS), are alpha-helical, amphipathic, hydrophobic and cationic and are active mainly against Gram-positive bacteria but inactive or weakly active against Gram-negative bacteria. Here, we report two novel members of the Temporin family, named Temporin-1Ee (FLINIAGVLSKLF(amide)) and Temporin-1 Re (FLPGLLAGLL(amide)), whose biosynthetic precursor structures were deduced from clones obtained from skin secretion-derived cDNA libraries of the European edible frog, Pelophylax kl. esculentus, by 'shotgun' cloning. Deduction of the molecular masses of each mature processed peptide from respective cloned cDNAs was used to locate respective molecules in reverse-phase HPLC fractions of secretion. Temporin-1Ee (MIC = 10 mu M) and Temporin-1Re (MIC = 60 mu M) were both found to be active against Gram-positive Staphylococcus aureus, but retaining a weak haemolytic activity. To our knowledge, Single-site substitutions can dramatically change the spectrum of activity of a given Temporin. Compared with Temporine-1Ec, just one chemically-conservative substitution (Val8 instead of Leu8), Temporin-1 Ee bearing a net charge of +2 displays broad-spectrum activity with particularly high potency on the clinically relevant Gram-negative strains, Escherichia coli (MIC = 40 mu M). These factors bode well for translating Temporins to be potential drug candidates for the design of new and valuable anti-infective agents. (C) 2016 Elsevier Inc. All rights reserved.
Michael J Conlon - One of the best experts on this subject based on the ideXlab platform.
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selection of antimicrobial frog peptides and Temporin 1dra analogs for treatment of bacterial infections based on their cytotoxicity and differential activity against pathogens
Chemical Biology & Drug Design, 2020Co-Authors: Rogier A Gaiser, Milena Mechkarska, Michael J Conlon, Jaione Ayerra Mangado, Wendy E Kaman, Peter Van Baarlen, Jerry M WellsAbstract:Cationic, amphipathic, α‐helical host‐defense peptides (HDPs) that are naturally secreted by certain species of frogs (Anura) possess potent broad‐spectrum antimicrobial activity and show therapeutic potential as alternatives to treat infections by multi‐drug resistant pathogens. Fourteen amphibian skin peptides and twelve analogues of Temporin‐1DRa were studied for their antimicrobial activities against clinically relevant human or animal skin infection‐associated pathogens. For comparison, antimicrobial potencies of frog skin peptides against a range of probiotic lactobacilli were determined. We used the VITEK 2 system to define a profile of antibiotic susceptibility for the bacterial panel. The minimal inhibitory concentration (MIC) values of the naturally occurring Temporin‐1DRa, CPF‐AM1, alyteserin‐1c, hymenochirin‐2B, and hymenochirin‐4B for pathogenic bacteria were 3‐ to 9‐fold lower than the values for the tested probiotic strains. Similarly, Temporin‐1DRa and its [Lys4], [Lys5] and [Aib8] analogues showed 5‐ to 6.5‐fold greater potency against the pathogens. In the case of PGLa‐AM1, XT‐7, Temporin‐1DRa and its [D‐Lys8] and [Aib13] analogues, no apoptosis or necrosis was detected in human peripheral blood mononuclear cells at concentrations below or above the MIC. Given the differential activity against commensal bacteria and pathogens, some of these peptides are promising candidates for further development into therapeutics for topical treatment of skin infections
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assessment of the potential of Temporin peptides from the frog rana temporaria ranidae as anti diabetic agents
Journal of Peptide Science, 2018Co-Authors: Vishal Musale, Yasser Abdelwahab, Peter R Flatt, Bruno Casciaro, Maria Luisa Mangoni, Michael J ConlonAbstract:Temporin A (FLPLIGRVLSGIL-NH2 ), Temporin F (FLPLIGKVLSGIL-NH2 ), and Temporin G (FFPVIGRILNGIL-NH2 ), first identified in skin secretions of the frog Rana temporaria, produced concentration-dependent stimulation of insulin release from BRIN-BD11 rat clonal β-cells at concentrations ≥1 nM, without cytotoxicity at concentrations up to 3 μM. Temporin A was the most effective. The mechanism of insulinotropic action did not involve an increase in intracellular Ca2+ concentrations. Temporins B, C, E, H, and K were either inactive or only weakly active. Temporins A, F, and G also produced a concentration-dependent stimulation of insulin release from 1.1B4 human-derived pancreatic β-cells, with Temporin G being the most potent and effective, and from isolated mouse islets. The data indicate that cationicity, hydrophobicity, and the angle subtended by the charged residues in the Temporin molecule are important determinants for in vitro insulinotropic activity. Temporin A and F (1 μM), but not Temporin G, protected BRIN-BD11 cells against cytokine-induced apoptosis (P < 0.001) and augmented (P < 0.001) proliferation of the cells to a similar extent as glucagon-like peptide-1. Intraperitoneal injection of Temporin G (75 nmol/kg body weight) together with a glucose load (18 mmol/kg body weight) in C57BL6 mice improved glucose tolerance with a concomitant increase in insulin secretion whereas Temporin A and F administration was without significant effect on plasma glucose levels. The study suggests that combination therapy involving agents developed from the Temporin A and G sequences may find application in Type 2 diabetes treatment.
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host defense peptides from lithobates forreri hylarana luctuosa and hylarana signata ranidae phylogenetic relationships inferred from primary structures of ranatuerin 2 and brevinin 2 peptides
Comparative Biochemistry and Physiology Part D: Genomics and Proteomics, 2014Co-Authors: Michael J Conlon, Milena Mechkarska, Per F. Nielsen, Laurent Coquet, Hubert Vaudry, Norbert Nowotny, Jolanta Kolodziejek, Thierry Jouenne, Jérôme Leprince, Jay D. KingAbstract:The primary structures of host-defense peptides present in frog skin secretions constitute useful molecular markers for establishing taxonomic classifications and investigating phylogenetic relationships between species within a particular genus. Peptidomic analysis has led to the characterization of multiple host-defense peptides in norepinephrine-stimulated skin secretions of three species of frogs from the family Ranidae: Lithobates forreri (Boulenger, 1883), Hylarana luctuosa (Peters, 1871), and Hylarana signata (Gunther, 1872). The L. forreri secretions contain ranatuerin-2 (2 peptides), brevinin-1 (4 peptides), and Temporin (1 peptide). The H. luctuosa secretions contain brevinin-2 (4 peptides), esculentin-1 (1 peptide), esculentin-2 (1 peptide), palustrin-2 (2 peptides), and Temporin (2 peptides). The H. signata secretions contain brevinin-2 (4 peptides), brevinin-1 (5 peptides), palustrin-2 (1 peptide), and Temporin (2 peptides). Cladistic analysis based upon the primary structures of 44 ranatuerin-2 peptides from 20 Lithobates species indicates a close phylogenetic relationship between L. forreri, Lithobates onca, and Lithobates yavapaiensis. A similar cladistic analysis based upon the primary structures of 27 brevinin-2 peptides from 8 Hylarana species provides support for a close phylogenetic relationship between H. signata and Hylarana picturata, while showing that the species are not conspecific, with H. luctuosa more distantly related.
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peptidomic analysis of skin secretions from the bullfrog lithobates catesbeianus ranidae identifies multiple peptides with potent insulin releasing activity
Peptides, 2011Co-Authors: Milena Mechkarska, Mohammed A Meetani, Jay D. King, Yasser Abdelwahab, Peter R Flatt, Laurent Coquet, Thierry Jouenne, Michael J ConlonAbstract:Using a combination of reversed-phase HPLC and electrospray mass spectrometry, peptidomic analysis of norepinephrine-stimulated skin secretions of the American bullfrog Litho bates catesbeianus Shaw, 1802 led to the identification and characterization of five newly described peptides (ranatuerin-1CBb, ranatuerin-2CBc, and -CBd, palustrin-2CBa, and Temporin-CBf) together with seven peptides previously isolated on the basis of their antimicrobial activity (ranatuerin-1CBa, ranatuerin-2CBa, brevinin-1CBa, and -1CBb, Temporin-CBa, -CBb, and -CBd). The abilities of the most abundant of the purified peptides to stimulate the release of insulin from the rat BRIN-BD11 clonal beta cell line were evaluated. Ranatuerin-2CBd (GFLDIIKNLGKTFAGHMLDKIRCTIGTCPPSP) was the most potent peptide producing a significant stimulation of insulin release (119% of basal rate, P<0.01) from BRIN-BD11 cells at a concentration of 30 nM, with a maximum response (236% of basal rate, P<0.001) at a concentration of 3 mu M. Ranatuerin-2CBd did not stimulate release of the cytosolic enzyme, lactate dehydrogenase at concentrations up to 3 mu M, indicating that the integrity of the plasma membrane had been preserved. Brevinin-1CBb (FLPFIAR-LAAKVFPSIICSVTKKC) produced the maximum stimulation of insulin release (285% of basal rate, P<0.001 at 3 mu M) but the peptide was cytotoxic at this concentration. (C) 2010 Elsevier Inc. All rights reserved.
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antimicrobial peptides from the skin secretions of the new world frogs lithobates capito and lithobates warszewitschii ranidae
Peptides, 2009Co-Authors: Michael J Conlon, Mohammed A Meetani, Laurent Coquet, Hubert Vaudry, Norbert Nowotny, Jolanta Kolodziejek, Thierry Jouenne, Jérôme Leprince, Jay D. KingAbstract:Abstract Taxonomic revisions within the anuran family Ranidae have established the genus Lithobates that currently comprises 49 species of frogs from the New World. Peptidomic analysis, using reversed-phase HPLC with on-line detection by electrospray mass spectrometry, has led to the identification of multiple antimicrobial peptides in norepinephrine-stimulated skin secretions of the North American frog Lithobates capito and the Central American frog Lithobates warszewitschii . Structural characterization of the peptides demonstrated that the L. capito secretions contained brevinin-1 (1), esculentin-1 (1), esculentin-2 (1), ranatuerin-2 (3), and Temporin (2) peptides. L. warszewitschii secretions contained brevinin-1 (1), esculentin-2 (1), ranatuerin-2 (2), and Temporin (1) peptides. Values in parentheses indicate number of peptides in each family. Temporin-CPa from L. capito , with the atypical structure IPPFIKKVLTTVF·NH 2 , also showed atypical growth-inhibitory activity having greater potency against Escherichia coli (MIC = 25 μM) and Candida albicans (MIC = 25 μM) than against Staphylococcus aureus (MIC = 50 μM). Phylogenetic analysis based upon the amino acid sequences of 37 ranatuerin-2 peptides from 17 species belonging to the genus Lithobates provides support for currently accepted taxonomic relationships. L. capito is sister-group to Lithobates sevosus in a clade that also contains Lithobates areolatus , and Lithobates palustris . L. warszewitschii is most closely related to the Central American species Lithobates tarahumarae and Lithobates vaillanti .
