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A M Fallon - One of the best experts on this subject based on the ideXlab platform.
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characterization of genomic dna encoding Cecropins from an aedes albopictus mosquito cell line
Insect Molecular Biology, 2002Co-Authors: A M FallonAbstract:We used cDNA probes from Aedes albopictus mosquito Cecropins AalCecA, B, and C to obtain genomic DNA copies and flanking DNA. Two gene copies (AalCecA1 and A2, AalCecB1 and B2, AalCecC1 and C2) encoding each of the three mature Cecropin peptides were recovered. All these genes had a similar organization, into two exons interrupted by a single short intron. AalCecA1 and AalCecA2 encode mature protein products that differ by one amino acid residue, while AalCecB1 and AalCecB2, AalCecC1 and AalCecC2 encode identical mature Cecropin peptides, respectively. The AalCecB and C gene pairs each share a common intergenic region of approximately 1 kb, with the two coding regions transcribed in opposite directions. With the exception of small insertions/deletions, the intergenic spacer region was highly conserved between the B1/C1 and B2/C2 clones. In transfected cells, 0.8 kb of upstream sequence was sufficient for inducible expression of AalCecA1. Within this region, a 28 bp sequence at positions –192 to –165 upstream of the transcription initiation site was found to contain a potential regulatory element. In electrophoretic mobility shift assays, synthetic double-stranded DNA containing this 28 bp sequence retarded protein in cytoplasmic and nuclear extracts from C7-10 cells.
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Cloning and expression of three Cecropin cDNAs from a mosquito cell line.
FEBS letters, 1999Co-Authors: Eric D Eccleston, A M FallonAbstract:We have characterized full-length cDNAs encoding three isoforms of the antibiotic Cecropin secreted by the C7-10 cell line from the mosquito, Aedes albopictus. The existence of two Cecropin isoforms that differed from the previously described AalCecA was predicted by mass spectrometry and amino acid sequence analysis of peptides that eluted from reversed phase high performance liquid chromatography as a single peak just behind the previously described Cecropin, AalCecA. Based on the amino acid sequence of the mature AalCecA peptide, we designed primers that amplified partial cDNAs encoding three different A. albopictus Cecropins in reverse transcriptase polymerase chain reactions. Rapid amplification of cDNA ends was then used to complete the cDNA sequences of AalCecA, AalCecB and AalCecC, respectively. Each cDNA encoded a translation product containing a signal peptide, a pro region, and a mature Cecropin peptide consistent with amino acid sequence data from chymotryptic digests. Although the mosquito Cecropins shared 70-86% identity among each other, they shared only approximately 40% identity to Cecropins from Drosophila melanogaster. Each of the Cecropins was expressed within 2 to 4 h after induction, and transcripts measuring 0.3 to 0.5 kb continued to accumulate over 24 h. The three Cecropins were secreted in roughly equimolar proportions, and 30 to 90% of AalCecB was amidated at the terminal glycine residue. In contrast, amidated forms of AalCecA and AalCecC constituted a smaller proportion of these isoforms.
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Cloning and expression of three Cecropin cDNAs from a mosquito cell line
FEBS Letters, 1999Co-Authors: Eric D Eccleston, A M FallonAbstract:We have characterized full-length cDNAs encoding three isoforms of the antibiotic Cecropin secreted by the C7–10 cell line from the mosquito, Aedes albopictus. The existence of two Cecropin isoforms that differed from the previously described AalCecA was predicted by mass spectrometry and amino acid sequence analysis of peptides that eluted from reversed phase high performance liquid chromatography as a single peak just behind the previously described Cecropin, AalCecA. Based on the amino acid sequence of the mature AalCecA peptide, we designed primers that amplified partial cDNAs encoding three different A. albopictus Cecropins in reverse transcriptase polymerase chain reactions. Rapid amplification of cDNA ends was then used to complete the cDNA sequences of AalCecA, AalCecB and AalCecC, respectively. Each cDNA encoded a translation product containing a signal peptide, a pro region, and a mature Cecropin peptide consistent with amino acid sequence data from chymotryptic digests. Although the mosquito Cecropins shared 70–86% identity among each other, they shared only ∼40% identity to Cecropins from Drosophila melanogaster. Each of the Cecropins was expressed within 2 to 4 h after induction, and transcripts measuring 0.3 to 0.5 kb continued to accumulate over 24 h. The three Cecropins were secreted in roughly equimolar proportions, and 30 to 90% of AalCecB was amidated at the terminal glycine residue. In contrast, amidated forms of AalCecA and AalCecC constituted a smaller proportion of these isoforms.
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Peptide Sequence of an Antibiotic Cecropin from the Vector Mosquito,Aedes albopictus
Biochemical and Biophysical Research Communications, 1998Co-Authors: Eric D Eccleston, A M FallonAbstract:Abstract We have identified a 35-amino acid antibiotic Cecropin secreted by an established mosquito cell line. C7-10 cells from the vector mosquito,Aedes albopictus,were incubated with heat-killedEscherichia coli,and materials secreted into the cell culture supernatant were recovered by acid precipitation. Following batch elution from Sep-Pak C18 cartridges and further purification by reverse phase high performance liquid chromatography (RP-HPLC) a predominant peak of antibacterial activity was characterized by mass spectrometry, amino acid composition analysis, and Edman degradation, yielding the sequence GGLKKLGKKLEGVGKRVFKASEKALPVAVGIKALG. Unlike other Cecropins, the peptide was not amidated at the C-terminus.Aedes albopictusCecropin A (AalCecA) is the first Cecropin to be described from a mosquito vector of human disease. Consistent with the classification of mosquitoes among the Dipteran suborder Nematocera, AalCecA shares only 36% amino acid identity with Cecropins fromDrosophila melanogasterand other Cyclorrhaphid flies, whose mature Cecropins share 80% to 100% amino acid identity.
Eric D Eccleston - One of the best experts on this subject based on the ideXlab platform.
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Cloning and expression of three Cecropin cDNAs from a mosquito cell line.
FEBS letters, 1999Co-Authors: Eric D Eccleston, A M FallonAbstract:We have characterized full-length cDNAs encoding three isoforms of the antibiotic Cecropin secreted by the C7-10 cell line from the mosquito, Aedes albopictus. The existence of two Cecropin isoforms that differed from the previously described AalCecA was predicted by mass spectrometry and amino acid sequence analysis of peptides that eluted from reversed phase high performance liquid chromatography as a single peak just behind the previously described Cecropin, AalCecA. Based on the amino acid sequence of the mature AalCecA peptide, we designed primers that amplified partial cDNAs encoding three different A. albopictus Cecropins in reverse transcriptase polymerase chain reactions. Rapid amplification of cDNA ends was then used to complete the cDNA sequences of AalCecA, AalCecB and AalCecC, respectively. Each cDNA encoded a translation product containing a signal peptide, a pro region, and a mature Cecropin peptide consistent with amino acid sequence data from chymotryptic digests. Although the mosquito Cecropins shared 70-86% identity among each other, they shared only approximately 40% identity to Cecropins from Drosophila melanogaster. Each of the Cecropins was expressed within 2 to 4 h after induction, and transcripts measuring 0.3 to 0.5 kb continued to accumulate over 24 h. The three Cecropins were secreted in roughly equimolar proportions, and 30 to 90% of AalCecB was amidated at the terminal glycine residue. In contrast, amidated forms of AalCecA and AalCecC constituted a smaller proportion of these isoforms.
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Cloning and expression of three Cecropin cDNAs from a mosquito cell line
FEBS Letters, 1999Co-Authors: Eric D Eccleston, A M FallonAbstract:We have characterized full-length cDNAs encoding three isoforms of the antibiotic Cecropin secreted by the C7–10 cell line from the mosquito, Aedes albopictus. The existence of two Cecropin isoforms that differed from the previously described AalCecA was predicted by mass spectrometry and amino acid sequence analysis of peptides that eluted from reversed phase high performance liquid chromatography as a single peak just behind the previously described Cecropin, AalCecA. Based on the amino acid sequence of the mature AalCecA peptide, we designed primers that amplified partial cDNAs encoding three different A. albopictus Cecropins in reverse transcriptase polymerase chain reactions. Rapid amplification of cDNA ends was then used to complete the cDNA sequences of AalCecA, AalCecB and AalCecC, respectively. Each cDNA encoded a translation product containing a signal peptide, a pro region, and a mature Cecropin peptide consistent with amino acid sequence data from chymotryptic digests. Although the mosquito Cecropins shared 70–86% identity among each other, they shared only ∼40% identity to Cecropins from Drosophila melanogaster. Each of the Cecropins was expressed within 2 to 4 h after induction, and transcripts measuring 0.3 to 0.5 kb continued to accumulate over 24 h. The three Cecropins were secreted in roughly equimolar proportions, and 30 to 90% of AalCecB was amidated at the terminal glycine residue. In contrast, amidated forms of AalCecA and AalCecC constituted a smaller proportion of these isoforms.
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Peptide Sequence of an Antibiotic Cecropin from the Vector Mosquito,Aedes albopictus
Biochemical and Biophysical Research Communications, 1998Co-Authors: Eric D Eccleston, A M FallonAbstract:Abstract We have identified a 35-amino acid antibiotic Cecropin secreted by an established mosquito cell line. C7-10 cells from the vector mosquito,Aedes albopictus,were incubated with heat-killedEscherichia coli,and materials secreted into the cell culture supernatant were recovered by acid precipitation. Following batch elution from Sep-Pak C18 cartridges and further purification by reverse phase high performance liquid chromatography (RP-HPLC) a predominant peak of antibacterial activity was characterized by mass spectrometry, amino acid composition analysis, and Edman degradation, yielding the sequence GGLKKLGKKLEGVGKRVFKASEKALPVAVGIKALG. Unlike other Cecropins, the peptide was not amidated at the C-terminus.Aedes albopictusCecropin A (AalCecA) is the first Cecropin to be described from a mosquito vector of human disease. Consistent with the classification of mosquitoes among the Dipteran suborder Nematocera, AalCecA shares only 36% amino acid identity with Cecropins fromDrosophila melanogasterand other Cyclorrhaphid flies, whose mature Cecropins share 80% to 100% amino acid identity.
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Peptide Sequence of an Antibiotic Cecropin from the Vector Mosquito,Aedes albopictus
Biochemical and Biophysical Research Communications, 1998Co-Authors: Eric D Eccleston, Ann M. FallonAbstract:Abstract We have identified a 35-amino acid antibiotic Cecropin secreted by an established mosquito cell line. C7-10 cells from the vector mosquito,Aedes albopictus,were incubated with heat-killedEscherichia coli,and materials secreted into the cell culture supernatant were recovered by acid precipitation. Following batch elution from Sep-Pak C18 cartridges and further purification by reverse phase high performance liquid chromatography (RP-HPLC) a predominant peak of antibacterial activity was characterized by mass spectrometry, amino acid composition analysis, and Edman degradation, yielding the sequence GGLKKLGKKLEGVGKRVFKASEKALPVAVGIKALG. Unlike other Cecropins, the peptide was not amidated at the C-terminus.Aedes albopictusCecropin A (AalCecA) is the first Cecropin to be described from a mosquito vector of human disease. Consistent with the classification of mosquitoes among the Dipteran suborder Nematocera, AalCecA shares only 36% amino acid identity with Cecropins fromDrosophila melanogasterand other Cyclorrhaphid flies, whose mature Cecropins share 80% to 100% amino acid identity.
Hans G. Boman - One of the best experts on this subject based on the ideXlab platform.
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DESIGN, SYNTHESIS AND ANTIBACTERIAL ACTIVITY OF Cecropin-LIKE MODEL PEPTIDES
International Journal of Peptide and Protein Research, 2009Co-Authors: Jürgen Fink, Anita Boman, Hans G. Boman, Robert B. MerrifieldAbstract:In order to investigate structure-activity relationships of Cecropins, model peptides that mimic certain structural features of the Cecropin molecules were designed and synthesized. The conformational analysis of Cecropins and the design of the model peptides were based on Chou-Fasman calculations. The peptides were synthesized by solid-phase methods and purified by reverse-phase liquid-chromatography on C18-silica columns. Their secondary structures were studied by circular dichroism measurements. Antibacterial activities against seven test organisms were determined and compared to the activities of the natural Cecropins A and B. These results were discussed on the basis of structural features of the model peptides and on model mechanisms. It was concluded that high antibacterial activity for this class of compounds requires a basic helical amphipathic N-terminal segment that is connected to a hydrophobic helical C-terminal segment by a flexible non-helical hinge region.
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antibacterial peptides designed as analogs or hybrids of Cecropins and melittin
International Journal of Peptide and Protein Research, 2009Co-Authors: D Wade, Anita Boman, A M V Silveira, S.a. Mitchell, Hans G. Boman, David Andreu, Robert B. MerrifieldAbstract:Eight new analogs of Cecropin A, two new analogs of melittin and 30 hybrid peptides containing sequences from Cecropins and melittin have been synthesized. The lengths of the peptides have varied from 37 residues (the length of Cecropin A) to 18 residues. The peptides have been assayed for lysis of sheep red blood cells and for antibacterial activity against two Gram negative and three Gram positive bacteria. The best analogs of Cecropin A maintained the anti-Escherichia coli activity of the parental peptide, and were not lytic for red blood cells. Melittin and its replacement analogs were all lytic for red blood cells, but an analog with transposed segments was not. Several of the hybrid peptides were found to be both non-hemolytic and highly active against all test bacteria. The data were used to define the structural requirements for antibacterial activity.
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sequence and specificity of two antibacterial proteins involved in insect immunity
Journal of Immunology, 2009Co-Authors: Håkan Steiner, Hans Bennich, Dan Hultmark, Ake Engstrom, Hans G. BomanAbstract:Immune responses have been described for many different insect species'. However, it is generally acknowledged that insects lack lymphocytes and immunoglobulins and their immune systems must therefore differ from those of vertebrates. An effective humoral immune response has been found in pupae of the cecropia moth, Hyalophora cecropia 1 . The expression of this multicomponent system requires de novo synthesis of RNA and proteins 2 and its broad antibacterial activity is due to at least three independent mechanisms 3 , the most well known of which is the insect lysozyme 4-7 . However, this enzyme is bactericidal for only a limited number of Gram-positive bacteria. We recently purified and characterized P9A and P9B, which are two small, basic proteins with potent antibacteral activity against Escherichia coli and several other Gram-negative bacteria'. We believe that P9A and P9B play an important part in the humoral immune responses described previously 8 and that the P9 proteins represent a new class of antibacterial agents for which we propose the name Cecropins. We describe here the primary structures of Cecropins A and B. We also show that Cecropin A is specific for bacteria in contrast to melittin, the main lytic component in bee venom 9 which lyses both bacteria and eukaryotic cells.
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the origin of Cecropins implications from synthetic peptides derived from ribosomal protein l1
FEBS Letters, 1999Co-Authors: Katrin Putsep, Staffan Normark, Hans G. BomanAbstract:We recently showed that Helicobacter pylori grown on plates produce Cecropin-like antibacterial peptides to which H. pylori is resistant. This antibacterial activity was traced to fragments from the N-terminus of ribosomal protein L1 (Putsep et al., Nature, April 22, 1999). The evolutionary suggestion from this finding has now been extended by the synthesis of eight peptides with sequences taken from the N-terminus of ribosomal protein L1 (RpL1) of five different species. Two peptides of different length derived from H. pylori RpL1 showed a potent antibacterial activity, while a peptide with the sequence from Escherichia coli was 20 times less active. Like Cecropins the H. pylori peptides were not cytolytic. We suggest that the Cecropins have evolved from ribosomal protein L1 of an ancestral intracellular pathogen that developed to a symbiont ending as an organelle. When the R1 gene moved into the host nucleus, a duplication provided a copy from which today Cecropins could have evolved.
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Interaction of the Mammalian Antibacterial Peptide Cecropin P1 with Phospholipid Vesicles
Biochemistry, 1995Co-Authors: Ehud Gazit, Anita Boman, Hans G. Boman, Yechiel ShaiAbstract:Cecropins are positively charged antibacterial polypeptides that were originally isolated from insects. Later on a mammalian homologue, Cecropin P1 (CecP), was isolated from pig intestines. While insect Cecropins are highly potent against both Gram-negative and Gram-positive bacteria, CecP is as active as insect Cecropins against Gram-negative but has reduced activity against Gram-positive bacteria. To gain insight into the mechanism of action of CecP and the molecular basis of its antibacterial specificity, the peptide and its proline incorporated analogue (at the conserved position found in insect Cecropins), P22-CecP, were synthesized and labeled on their N-terminal amino-acids with fluorescent probes, without significantly affecting their antibacterial activities. Fluorescence studies indicated that the N-terminal of CecP is located on the surface of phospholipid membranes. Binding experiments revealed that CecP binds acidic phosphatidylserine/phosphatidylcholine (PS/PC) vesicles better than zwitterionic PC vesicles, which correlates with its ability to permeate the former better than the latter. The shape of the binding isotherms suggest that CecP, like insect Cecropin, binds phospholipids in a simple, noncooperative manner. However, resonance energy transfer (RET) measurements revealed that, unlike insect Cecropins, CecP does not aggregate in the membrane even at relatively high peptide to lipid ratios. The stoichiometry of CecP binding to vesicles suggests that amount of CecP sufficient to form a monolayer causes vesicle permeation. In spite of the incorporation of the conserved proline in P22-CecP, the analogue has reduced antibacterial activity, which correlates with its reduced alpha-helical structure and its lower partitioning and membrane permeating activity with phospholipid vesicles. Taken together, our results support a mechanism in which CecP disrupts the structure of the bacterial membrane by (i) binding of peptide monomers to the acidic surface of the bacterial membrane and (ii) disintegrating the bacterial membrane by disrupting the lipid packing in the bilayers. These results, combined with data reported for other antibacterial polypeptides, suggest that the organization of peptide monomers within phospholipid membranes contributes to Gram-positive/Gram-negative antibacterial specificity.
Thomas T. Chen - One of the best experts on this subject based on the ideXlab platform.
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RNA-Seq analysis of differentially expressed genes relevant to innate and adaptive immunity in Cecropin P1 transgenic rainbow trout (Oncorhynchus mykiss)
BMC, 2018Co-Authors: Yueh-chiang Han, Chun-mean Lin, Thomas T. ChenAbstract:Abstract Background In the past years, our laboratory successfully generated transgenic rainbow trout bearing Cecropin P1 transgene. These fish exhibited resistant characteristic to infection by Aeromonas salmonicida, Infectious Hematopoietic Necrosis Virus (IHNV) and Ceratomyxa shasta (a parasitic pathogen). Previously, treating rainbow trout macrophage cells (RTS-11) with Cecropin B, pleurocidin and CF17, respectively, resulted in elevated expression of two pro-inflammatory genes, e.g. cyclooxygenase-2 (cox-2) and interleukin-1β (il-1β). In addition, a profiling of global gene expression by 44 k salmonid microarray analysis was conducted, and the results showed that immune relevant processes have been perturbed in cecopin P1 transgenic rainbow trout. Therefore, we hypothesized that Cecropin P1 may not only eliminate pathogens directly, but also modulate the host immune systems, leading to increased resistance against pathogen infections. To confirm this hypothesis, we performed de novo mRNA deep sequencing (RNA-Seq) to analyze the transcriptomic expression profiles in three immune competent tissues of Cecropin P1 transgenic rainbow trout. Results De novo sequencing of mRNA of the rainbow trout spleen, liver and kidney tissues were conducted by second-generation Illumina system, followed by Trinity assembly. Tissue specific unigenes were obtained, and annotated according to the Gene Ontology (GO) and the Nucleotide Basic Local Alignment Search Tool (BLAST). Over 2000 differentially expressed genes (DEGs) were determined by normalized ratio of Reads Per Kilobase of transcript per million mapped reads (RPKM) among the transgenic and non-transgenic fish in a tissue specific manner, and there were 82 DEGs in common among the three tissues. In addition, the enrichment analysis according to Gene Ontology Biological Process (GO:BP), and Kyoto Encyclopedia of Genes and Genomes (KEGG) based pathway analysis associated with innate/adaptive immunity of fish were also performed to illustrate the altered immune-related functions in each tissue. Conclusions According to the RNA-Seq data, the correlations between alteration of gene expression profiles and the functional perturbations of the host immune processes were revealed. In comparison with the results of cDNA microarray analysis conducted by Lo et al., the overall results supported our hypothesis that the gene product of Cecropin P1 transgene may not only directly eliminate pathogens, but also modulate the host immune system. Results of this study present valuable genetic information for Oncorhynchus mykiss, and will benefit future studies on the immunology of this fish species
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Inducible Resistance of Fish Bacterial Pathogens to the Antimicrobial Peptide Cecropin B
Antimicrobial Agents and Chemotherapy, 2008Co-Authors: Ulysses W Sallum, Thomas T. ChenAbstract:Cecropin B is a cationic antimicrobial peptide originally isolated from the diapausing pupae of the giant silk moth, Hylphora cecropia. Cecropin B elicits its antimicrobial effects through disruption of the anionic cell membranes of gram-negative bacteria. Previous work by our laboratory demonstrated that a constitutively expressed Cecropin B transgene conferred enhanced resistance to bacterial infection in medaka. The development of antibiotic resistance by pathogenic bacteria is a growing problem. The potential for fish bacterial pathogens to develop resistance to Cecropin B was addressed in this study. Four fish bacterial pathogens were selected for the study based on their importance in aquaculture. Vibrio anguillarum, Vibrio vulnificus, and Yersinia ruckeri all exhibited inducible resistance to Cecropin B. The inducible resistance of these three pathogens was correlated with reversible changes in their ultrastructures, as observed by scanning electron microscopy. V. anguillarum was demonstrated to become more adhesive to a CHSE-214 cell monolayer and to cause increased cumulative mortality in medaka following exposure to Cecropin B. This work demonstrates that the resistance of fish bacterial pathogens to Cecropin B is inducible and suggests that resistance to other cationic antimicrobial peptides may occur through similar means. The observed changes in ultrastructure and infectivity suggest that resistance to antimicrobial peptides is an integral part of the pathogenesis of fish gram-negative bacterial pathogens.
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Inducible Resistance of Fish Bacterial Pathogens to the Antimicrobial Peptide Cecropin B
Antimicrobial Agents and Chemotherapy, 2008Co-Authors: Ulysses W Sallum, Thomas T. ChenAbstract:Cecropin B is a cationic antimicrobial peptide originally isolated from the diapausing pupae of the giant silk moth, Hylphora cecropia. Cecropin B elicits its antimicrobial effects through disruption of the anionic cell membranes of gram-negative bacteria. Previous work by our laboratory demonstrated that a constitutively expressed Cecropin B transgene conferred enhanced resistance to bacterial infection in medaka. The development of antibiotic resistance by pathogenic bacteria is a growing problem. The potential for fish bacterial pathogens to develop resistance to Cecropin B was addressed in this study. Four fish bacterial pathogens were selected for the study based on their importance in aquaculture. Vibrio anguillarum, Vibrio vulnificus, and Yersinia ruckeri all exhibited inducible resistance to Cecropin B. The inducible resistance of these three pathogens was correlated with reversible changes in their ultrastructures, as observed by scanning electron microscopy. V. anguillarum was demonstrated to become more adhesive to a CHSE-214 cell monolayer and to cause increased cumulative mortality in medaka following exposure to Cecropin B. This work demonstrates that the resistance of fish bacterial pathogens to Cecropin B is inducible and suggests that resistance to other cationic antimicrobial peptides may occur through similar means. The observed changes in ultrastructure and infectivity suggest that resistance to antimicrobial peptides is an integral part of the pathogenesis of fish gram-negative bacterial pathogens.
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Bactericidal activity of Cecropin B and Cecropin P1 expressed in fish cells (CHSE-214): application in controlling fish bacterial pathogens
Aquaculture, 2003Co-Authors: Aliye Sarmasik, Thomas T. ChenAbstract:Abstract Cecropins are a group of antimicrobial peptides which have bactericidal activity against a broad range of bacteria. To date, the Cecropins used in a variety of studies were either purified from their natural source or obtained by chemical synthesis. The present study was conducted to test whether bactericidally active Cecropins could be expressed in a fish cell line. For this purpose, Chinook salmon embryo cells (CHSE-214) were transfected with Cecropin transgene constructs: Hyalophora cecropia preproCecropin B, proCecropin B, Cecropin B, and porcine P 1 Cecropin. From the transfected cells, single cell clones were selected and screened for the presence of Cecropin gene constructs by PCR amplification. The expression of the Cecropin transgene in the PCR positive clones was determined by RT-PCR reaction. Southern blot hybridization results showed that the Cecropin gene constructs were integrated into the genome in a multiple integration pattern. Bactericidal activity of the Cecropins, synthesized from transgene constructs, was detected using inhibition zone assay for fish pathogenic bacteria: Aeromonas hydrophila , Pseudomonas fluorescens , and Vibrio anguillarum . Cecropin antimicrobial peptides produced in CHSE-214 cells possess bactericidal activity against these three fish pathogenic bacteria.
Philippe Bulet - One of the best experts on this subject based on the ideXlab platform.
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Solution structures of stomoxyn and spinigerin, two insect antimicrobial peptides with an alpha-helical conformation
Biopolymers, 2006Co-Authors: Céline Landon, Philippe Bulet, H. Meudal, Nathalie Boulanger, Françoise VovelleAbstract:Stomoxyn and spinigerin belong to the class of linear cysteine-free insect antimicrobial peptides that kill a range of microorganisms, parasites, and some viruses but without any lytic activity against mammalian erythrocytes. Stomoxyn is localized in the gut epithelium of the nonvector stable fly that is sympatric with the trypanosome vector tsetse fly. Spinigerin is stored and secreted by hemocytes from the fungus-growing termite. The structure of synthetic stomoxyn and spinigerin in aqueous solution and in TFE/water mixtures was analyzed by CD and NMR spectroscopy combined with molecular modeling calculations. Stomoxyn and spinigerin adopt a flexible random coil structure in water while both assume a stable helical structure in the presence of TFE. In 50% TFE, the structure of stomoxyn is typical of Cecropins, including an amphipathic helix at the N-terminus and a hydrophobic C-terminus with helical features that probably fold in a helical conformation at higher TFE concentration. In contrast to stomoxyn, spinigerin acquires very rapidly a helical conformation. In 10% TFE the helix is highly bent and the structure is poorly defined. In 50% TFE, the helical structure is well defined all along its sequence, and the slightly bent -helix displays an amphiphilic character, as observed for magainin 2. The structural similarities between stomoxyn and Cecropin A from Hyalophora cecropia and between spinigerin and magainin 2 suggest a similar mode of action on the bacterial membranes of both pairs of peptides. Our results also confirm that TFE induces helix formation and propagation for amino acids showing helical propensity in water but also enhances the helix propagation propensity of nonpolar -branched residues.
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cloning and analysis of a Cecropin gene from the malaria vector mosquito anopheles gambiae
Insect Molecular Biology, 2000Co-Authors: Jacopo Vizioli, Jules A Hoffmann, Philippe Bulet, Maurice Charlet, Carl Lowenberger, Claudia Blass, H M Muller, George Dimopoulos, F C Kafatos, Adam M RichmanAbstract:Parasites of the genus Plasmodium are transmitted to mammalian hosts by anopheline mosquitoes. Within the insect vector, parasite growth and development are potentially limited by antimicrobial defence molecules. Here, we describe the isolation of cDNA and genomic clones encoding a Cecropin antibacterial peptide from the malaria vector mosquito Anopheles gambiae. The locus was mapped to polytene division 1C of the X chromosome. Cecropin RNA was induced by infection with bacteria and Plasmodium. RNA levels varied in different body parts of the adult mosquito. During development, Cecropin expression was limited to the early pupal stage. The peptide was purified from both adult mosquitoes and cell culture supernatants. Anopheles gambiae synthetic Cecropins displayed activity against Gram-negative and Gram-positive bacteria, filamentous fungi and yeasts.
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antimicrobial activity spectrum cdna cloning and mrna expression of a newly isolated member of the Cecropin family from the mosquito vector aedes aegypti
Journal of Biological Chemistry, 1999Co-Authors: Carl Lowenberger, Jacopo Vizioli, Maurice Charlet, Adam M Richman, Sofie Kamal, Bruce M Christensen, Philippe BuletAbstract:Abstract An antimicrobial peptide belonging to the Cecropin family was isolated from the hemolymph of bacteria-challenged adult Aedes aegypti. This new peptide, named Cecropin A, was purified to homogeneity and fully characterized after cDNA cloning. The 34-residue A. aegypti Cecropin A is different from the majority of reported insect Cecropins in that it is devoid of a tryptophan residue and C-terminal amidation. The importance of these two structural features on the activity spectrum was investigated using a chemically synthesized peptide. A comparison of the antimicrobial activity spectrum of A. aegypti and DrosophilaCecropin A showed a lower activity for the mosquito molecule. A. aegypti Cecropin mRNA expression was not detected by Northern blot or reverse transcription-polymerase chain reaction analysis in any immature stage of the mosquito, nor in naive adults, but it was observed in challenged adults 6 h after bacteria inoculation, and it continued over 7–10 days.