RESUMO
Natural radiation of geological origin is a common phenomenon in Brazil, a country where radioactive agents such as uranium may be often found. As an unstable atom, uranium undergoes radioactive decay with the generation of a series of decay by-products, including radon, which may be highly genotoxic and trigger several pathological processes, among which cancer. Because it is a gas, radon may move freely between cracks and gaps in the ground, seeping upwards into the buildings and in the environment. In this study, two Drosophila melanogaster Meigen (Diptera, Drosophilidae) strains called Oregon-R and Wild (collected in a non-radioactive environment) were exposed to atmospheric radiation in the Lajes Pintadas city, in the semiarid zone of northeastern Brazil. After six days of environmental exposure, the organisms presented genetic damage significantly higher than that of the negative control group. The genotoxic effects observed reinforce the findings of other studies carried out in the same region, which warn about the environmental risks related to natural radioactivity occurrence. The results also validate the use of the Comet assay in hemocytes of D. melanogaster as a sensitive test to detect genotoxicity caused by natural radiation, and the use of a recently collected D. melanogaster strain in the environmental of radon.
Assuntos
Radiação de Fundo/efeitos adversos , Ensaio Cometa/métodos , Dano ao DNA , Drosophila melanogaster/efeitos da radiação , Exposição Ambiental , Radônio/toxicidade , Animais , Brasil , Clima Desértico , Drosophila melanogaster/genética , Hemócitos/química , Sensibilidade e Especificidade , Urânio/toxicidadeRESUMO
Clavanins is a class of peptides (23aa) histidine-rich, free of post-translational modifications. Clavanins have been studied largely for their ability to disrupt bacterial membranes. In the present study, the interaction of clavanin A with membranes was assessed by dynamic light scattering, zeta potential and permeabilization assays. We observed through those assays that clavanin A lysis bacterial cells at concentrations corresponding to its MIC. Further, the structure and function of clavanin A was investigated. To better understand how clavanin interacted with bacteria, its NMR structure was elucidated. The solution state NMR structure of clavanin A in the presence of TFE-d3 indicated an α-helical conformation. Secondary structures, based on circular dichroism measurements in anionic sodium dodecyl sulfate (SDS) and TFE (2,2,2-trifluorethanol), in silico lipid-peptide docking and molecular simulations with lipids DPPC and DOPC revealed that clavanin A can adopt a variety of folds, possibly influencing its different functions. Microcalorimetry assays revealed that clavanin A was capable of discriminating between different lipids. Finally, clavanin A was found to eradicate bacterial biofilms representing a previously unrecognized function.
Assuntos
Bactérias/efeitos dos fármacos , Biofilmes/efeitos dos fármacos , Proteínas Sanguíneas/química , Bicamadas Lipídicas/metabolismo , Urocordados/metabolismo , Animais , Fenômenos Fisiológicos Bacterianos/efeitos dos fármacos , Proteínas Sanguíneas/farmacologia , Membrana Celular/efeitos dos fármacos , Dicroísmo Circular , Difusão Dinâmica da Luz , Hemócitos/química , Hemócitos/metabolismo , Testes de Sensibilidade Microbiana , Simulação de Acoplamento Molecular , Estrutura Secundária de Proteína , Urocordados/químicaRESUMO
The melanization reaction promoted by the prophenoloxidase-activating system is an essential defense response in invertebrates subjected to regulatory mechanisms that are still not fully understood. We report here the finding and characterization of a novel trypsin inhibitor, named panulirin, isolated from the hemocytes of the spiny lobster Panulirus argus with regulatory functions on the melanization cascade. Panulirin is a cationic peptide (pI 9.5) composed of 48 amino acid residues (5.3 kDa), with six cysteine residues forming disulfide bridges. Its primary sequence was determined by combining Edman degradation/N-terminal sequencing and electrospray ionization-MS/MS spectrometry. The low amino acid sequence similarity with known proteins indicates that it represents a new family of peptidase inhibitors. Panulirin is a competitive and reversible tight-binding inhibitor of trypsin (Ki = 8.6 nm) with a notable specificity because it does not inhibit serine peptidases such as subtilisin, elastase, chymotrypsin, thrombin, and plasmin. The removal of panulirin from the lobster hemocyte lysate leads to an increase in phenoloxidase response to LPS. Likewise, the addition of increasing concentrations of panulirin to a lobster hemocyte lysate, previously depleted of trypsin-inhibitory activity, decreased the phenoloxidase response to LPS in a concentration-dependent fashion. These results indicate that panulirin is implicated in the regulation of the melanization cascade in P. argus by inhibiting peptidase(s) in the pathway toward the activation of the prophenoloxidase enzyme.
Assuntos
Proteínas de Artrópodes/metabolismo , Catecol Oxidase/metabolismo , Precursores Enzimáticos/metabolismo , Palinuridae/metabolismo , Inibidores da Tripsina/metabolismo , Sequência de Aminoácidos , Animais , Proteínas de Artrópodes/química , Proteínas de Artrópodes/genética , Catecol Oxidase/química , Catecol Oxidase/genética , Relação Dose-Resposta a Droga , Ativação Enzimática/efeitos dos fármacos , Ativação Enzimática/fisiologia , Precursores Enzimáticos/química , Precursores Enzimáticos/genética , Hemócitos/química , Hemócitos/citologia , Hemócitos/metabolismo , Lipopolissacarídeos/farmacologia , Dados de Sequência Molecular , Palinuridae/química , Palinuridae/genética , Tripsina/química , Inibidores da Tripsina/química , Inibidores da Tripsina/genéticaRESUMO
This study presents the morphological description and histochemical characterization of gill filaments of the Brazilian endemic bivalve Diplodon expansus, aiming to broaden the morphological knowledge of this species and establish the structure of the gills that will serve as control in histopathological studies applied to biomonitoring. The gill filaments are divided into three zones: frontal, intermediate, and abfrontal. In the center of the filament, haemocytes circulate through the haemolymph vessel, which is internally lined by endothelium. The frontal surface of the filament is covered with cilia, the lateral surface exhibits aquifer ducts, and the abfrontal surface presents ciliated and nonciliated cells. The epithelium of the filaments is composed of ciliated cells, nonciliated absorptive cells, and mucocytes. The support of the filaments is made by two specialized structures called skeletal rod and skeletal loop. Based on the obtained information, the gill filaments of the studied species present some peculiar characteristics that are not yet reported in detail in the literature such as the simultaneous presence of skeletal rod and skeletal loop. On the other hand, the general constitution of the filament is similar to that described for both marine and limnic bivalves and seems to be suitable for ecotoxicological studies.
Assuntos
Bivalves/anatomia & histologia , Bivalves/química , Brânquias/ultraestrutura , Histocitoquímica/métodos , Animais , Brasil , Cílios/química , Colágeno/química , Células Endoteliais/química , Epitélio/química , Brânquias/química , Hemócitos/química , Hemolinfa/química , Microscopia Eletrônica de Varredura , Especificidade da EspécieRESUMO
Most experimental procedures on molluscs are done after acclimatization of wild animals to lab conditions. Similarly, short-term acclimation is often unavoidable in a field survey when biological analysis cannot be done within the day of sample collection. However, acclimatization can affect the general physiological condition and particularly the immune cell responses of molluscs. Our aim was to study the changes in the hemocyte characteristics of the Pacific oyster Crassostrea gigas and the carpet shell clam Ruditapes decussatus acclimated 1 or 2 days under emersed conditions at 14 ± 1 °C and for 1, 2, 7, or 10 days to flowing seawater conditions (submerged) at 9 ± 1 °C, when compared to hemolymph withdrawn from organisms sampled in the field and immediately analyzed in the laboratory (unacclimated). The hemocyte characteristics assessed by flow cytometry were the total (THC) and differential hemocyte count, percentage of dead cells, phagocytosis, and reactive oxygen species (ROS) production. Dead hemocytes were lower in oysters acclimated both in emersed and submerged conditions (1%-5%) compared to those sampled in the field (7%). Compared to oysters, the percentage of dead hemocytes was lower in clams (0.4% vs. 1.1%) and showed a tendency to decrease during acclimatization in both emersed and submerged conditions. In comparison to organisms not acclimated, the phagocytosis of hemocytes decreased in both oysters and clams acclimated under submerged conditions, but was similar in those acclimated in emersed conditions. The ROS production remained stable in both oysters and clams acclimated in emersed conditions, whereas in submerged conditions ROS production did not change in both the hyalinocytes and granulocytes of oysters, but increased in clams. In oysters, the THC decreased when they were acclimated 1 and 2 days in submerged conditions and was mainly caused by a decrease in granulocytes, but the decrease in THC in oysters acclimated 2 days in emersed conditions was caused by a decrease in hyalinocytes and small agranular cells. In clams, the THC was significantly lower in comparison to those not acclimated, regardless of the conditions of the acclimatization. These findings demonstrate that hemocyte characteristics were differentially affected in both species by the tested conditions of acclimatization. The phagocytosis and ROS production in clams and phagocytosis in oysters were not different in those acclimated for 1 day under both conditions, i.e. emersed and submerged, and those sampled in the field (unacclimated). The THC was significantly affected by acclimatization conditions, so the differences between clams and oysters should be considered in studies where important concentrations of hemocytes are required. The difference in the immune response between both species could be related to their habitat (epifaunal vs. infaunal) and their ability of resilience to manipulation and adaptation to captivity. Our results suggest that functional characteristics of hemocytes should be analyzed in both oysters and clams during the first 1 or 2 days, preferably acclimated under emersed rather than submerged conditions.
Assuntos
Aclimatação/fisiologia , Bivalves/citologia , Crassostrea/citologia , Hemócitos/fisiologia , Animais , Bivalves/fisiologia , Contagem de Células Sanguíneas/veterinária , Crassostrea/fisiologia , Citometria de Fluxo , Hemócitos/química , Fagocitose/fisiologia , Espécies Reativas de Oxigênio/metabolismo , Especificidade da EspécieRESUMO
This study aimed to verify if Dinophysis acuminata natural blooms affected the immune system of three bivalves: the oyster, Crassostrea gigas, the mussel, Perna perna, and the clam, Anomalocardia brasiliana. Animals were obtained from a renowned mariculture farm in the southern bay of Santa Catarina Island during, and 30 days after (controls), an algal bloom. Various immunological parameters were assessed in the hemolymph of the animals: total and differential hemocyte counts, percentage of apoptotic hemocytes, protein concentration, hemagglutinating titer and phenoloxidase activity. The results showed that the mussel was the most affected species, with several altered immune parameters, whereas the immunological profile of clams and oysters was partially and completely unaffected, respectively.
Assuntos
Bivalves/imunologia , Crassostrea/imunologia , Dinoflagellida/crescimento & desenvolvimento , Eutrofização , Perna (Organismo)/imunologia , Animais , Apoptose , Bivalves/classificação , Brasil , Crassostrea/classificação , Dinoflagellida/patogenicidade , Hemaglutinação , Hemócitos/química , Hemócitos/citologia , Hemolinfa/química , Ilhas , Monofenol Mono-Oxigenase/análise , Monofenol Mono-Oxigenase/metabolismo , Ácido Okadáico/análise , Perna (Organismo)/classificaçãoRESUMO
In this work we evaluated the ability of different types of antimicrobial peptides to promote permeabilization and growth inhibition of Acanthamoeba castellanii trophozoites, which cause eye keratitis. We used cationic alpha-helical peptides P5 and P6, corresponding to the N-terminus of the pore-forming protein from Triatoma infestans, a blood-sucking insect, and a beta-hairpin amphipathic molecule (gomesin), of the spider Acanthoscurria gomesiana haemocytes. A. castellanii permeabilization was obtained after 1 h incubation with micromolar concentrations of both types of peptides. While permeabilization induced by gomesin increased with longer incubations, P5 permeabilization did not increase with time and occurred at doses that are more toxic for SIRC cells. P5, however, at doses below the critical dose used to kill rabbit corneal cells was quite effective in promoting growth inhibition. Similarly, P5 was more effective when serine protease inhibitor was added simultaneously to the permeabilization assay. High performance chromatography followed by mass spectrometry analysis confirmed that, in contrast to gomesin, P5 is hydrolysed by A. castellanii culture supernatants. We conclude that the use of antimicrobial peptides to treat A. castellanii infections requires the search of more specific peptides that are resistant to proteolysis.
Assuntos
Acanthamoeba castellanii/efeitos dos fármacos , Peptídeos Catiônicos Antimicrobianos/farmacologia , Proteínas e Peptídeos Salivares/farmacologia , Ceratite por Acanthamoeba/tratamento farmacológico , Acanthamoeba castellanii/crescimento & desenvolvimento , Acanthamoeba castellanii/metabolismo , Animais , Peptídeos Catiônicos Antimicrobianos/síntese química , Aracnídeos/química , Linhagem Celular , Permeabilidade da Membrana Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Hemócitos/química , Estrutura Secundária de Proteína , Coelhos , Proteínas e Peptídeos Salivares/síntese química , Proteínas e Peptídeos Salivares/química , Inibidores de Serina Proteinase/farmacologia , Triatoma/química , Trofozoítos/efeitos dos fármacos , Trofozoítos/metabolismoRESUMO
Ki-67 is a protein expressed in the nucleus of several species during cell-division, being absent during the GO resting phase of the cellular cycle. During attempts to disclose mitosis in the so-called " amebocyte-producing organ " in Biomphalaria glabrata infected with Schistosoma mansoni, the parasite multiplying forms appeared strongly marked for Ki-67, while the snail tissues were completely negative. These data are worth registering to complement general data on Ki-67, and to help future studies on the relationship of the parasite and of its intermediate host.
Assuntos
Biomphalaria/citologia , Antígeno Ki-67/metabolismo , Schistosoma mansoni/fisiologia , Animais , Biomphalaria/parasitologia , Hemócitos/química , Interações Hospedeiro-Parasita/fisiologia , Imuno-Histoquímica , Camundongos , Índice Mitótico , Coloração e RotulagemRESUMO
Crustins are antimicrobial peptides initially identified in the hemocytes of the crab Carcinus maenas (11.5-kDa peptide or carcinin) and recently also recognized in penaeid shrimps and other crustacean species. The aim of this study was to identify sequences encoding for crustins from the hemocytes of four Brazilian penaeid species: Farfantepenaeus paulensis, Farfantepenaeus subtilis, Farfantepenaeus brasiliensis and Litopenaeus schmitti. Using primers based on consensus nucleotide alignment of crustins from different crustaceans, cDNA sequences coding for crustins in all indigenous penaeid species were amplified. The obtained four crustin sequences encoded for peptides containing a hydrophobic N-terminal region rich in glycine repeats and a C-terminal part with 12 cysteine residues and a conserved whey acidic protein domain. All obtained crustin sequences showed high amino acidic similarity among each other and with crustins from litopenaeid shrimps (76-98%). This is the first report of crustins in native Brazilian penaeid shrimps.
Assuntos
Peptídeos Catiônicos Antimicrobianos/genética , Hemócitos/química , Penaeidae/citologia , Peptídeos/genética , Sequência de Aminoácidos , Animais , Clonagem Molecular , DNA Complementar/metabolismo , Dados de Sequência Molecular , Peptídeos/fisiologia , Homologia de Sequência de AminoácidosRESUMO
Ki-67 is a protein expressed in the nucleus of several species during cell-division, being absent during the GO resting phase of the cellular cycle. During attempts to disclose mitosis in the so-called " amebocyte-producing organ " in Biomphalaria glabrata infected with Schistosoma mansoni, the parasite multiplying forms appeared strongly marked for Ki-67, while the snail tissues were completely negative. These data are worth registering to complement general data on Ki-67, and to help future studies on the relationship of the parasite and of its intermediate host.
Assuntos
Animais , Camundongos , Biomphalaria/citologia , /metabolismo , Schistosoma mansoni/fisiologia , Biomphalaria/parasitologia , Hemócitos/química , Interações Hospedeiro-Parasita/fisiologia , Imuno-Histoquímica , Índice Mitótico , Coloração e RotulagemRESUMO
We have isolated a 417Da antibacterial molecule, named mygalin, from the hemocytes of the spider Acanthoscurria gomesiana. The structure of mygalin was elucidated by tandem mass spectrometry (MS/MS) and by two spectroscopic techniques, nuclear magnetic resonance (NMR) and ultraviolet (UV) spectroscopy. Mygalin was identified as bis-acylpolyamine N1,N8-bis(2,5-dihydroxybenzoyl)spermidine, in which the primary amino groups of the spermidine are acylated with the carboxyl group of the 2,5-dihydroxybenzoic acid. Mygalin was active against Escherichia coli at 85muM, being this activity inhibited completely by catalase. Therefore, the antibacterial activity of mygalin was attributed to its production of hydrogen peroxide (H(2)O(2)). The putative mechanisms of formation of H(2)O(2) from mygalin are discussed. To our knowledge this is the first report of one bis-acylpolyamine with antibacterial activity purified from animal source.
Assuntos
Antibacterianos/química , Antibacterianos/farmacologia , Hemócitos/química , Espermidina/análogos & derivados , Aranhas/química , Animais , Antibacterianos/isolamento & purificação , Candida albicans/efeitos dos fármacos , Cromatografia em Gel , Cromatografia Líquida de Alta Pressão , Escherichia coli/efeitos dos fármacos , Isomerismo , Espectroscopia de Ressonância Magnética , Espectrometria de Massas , Micrococcus luteus/efeitos dos fármacos , Estrutura Molecular , Espermidina/química , Espermidina/isolamento & purificação , Espermidina/farmacologiaRESUMO
Lectins/carbohydrate binding can be involved in the Schistosoma mansoni recognition and activation of the Biomphalaria hemocytes. Therefore, expression of lectin ligands on Biomphalaria hemocytes would be associated with snail resistance against S. mansoni infection. To test this hypothesis, circulating hemocytes were isolated from B. glabrata BH (snail strain highy susceptible to S. mansoni), B. tenagophila Cabo Frio (moderate susceptibility), and B. tenagophila Taim (completely resistant strains), labelled with FITC conjugated lectins (ConA, PNA, SBA, and WGA) and analyzed under fluorescence microscopy. The results demonstrated that although lectin-labelled hemocytes were detected in hemolymph of all snail species tested, circulating hemocytes from both strains of B. tenagophila showed a larger number of lectin-labelled cells than B. glabrata. Moreover, most of circulating hemocytes of B. tenagophila were intensively labelled by lectins PNA-FITC and WGA-FITC, while in B. glabrata small hemocytes were labeled mainly by ConA. Upon S. mansoni infection, lectin-labelled hemocytes almost disappeared from the hemolymph of Taim and accumulated in B. glabrata BH. The role of lectins/carbohydrate binding in resistance of B. tengophila infection to S. mansoni is still not fully understood, but the data suggest that there may be a correlation to its presence with susceptibility or resistance to the parasite.
Assuntos
Animais , Biomphalaria/parasitologia , Hemócitos/química , Lectinas/metabolismo , Schistosoma mansoni/fisiologia , Biomphalaria/classificação , Contagem de Células , Interações Hospedeiro-Parasita , Microscopia de Fluorescência , FagocitoseRESUMO
Ergosan an algal product containing 1% alginic acid, developed for use in aquaculture and reported to have immunomodulatory activity, was administered orally to intermoult adult white shrimp (Litopenaeus vannamei) for 15 days. Examination of haemolymph proteins using SDS-PAGE did not reveal any obvious differences between control and Ergosan treated shrimp. Similarly, total haemocyte counts were found to be roughly equivalent for both the control and experimental samples. However, differential analysis of haemocyte populations revealed marked changes in terms of the relative levels of hyaline, semi-granular, and particularly granular haemocytes between the two groups. Moreover, enhancement of the in vitro antimicrobial activity of haemolymph towards two shrimp pathogenic Vibrio isolates was recorded for shrimp fed with Ergosan. Finally, shrimp fed with Ergosan showed a significant increase in relative growth when compared with control groups.
Assuntos
Alginatos/administração & dosagem , Decápodes/fisiologia , Hemócitos/química , Hemolinfa/química , Imunidade Inata/efeitos dos fármacos , Animais , Antibacterianos , Decápodes/efeitos dos fármacos , Ácido Glucurônico/administração & dosagem , Hemolinfa/fisiologia , Ácidos Hexurônicos/administração & dosagem , Imunização , Vibrio/efeitos dos fármacos , Vibrio/crescimento & desenvolvimento , Vibrio parahaemolyticusRESUMO
The presence of an effective immune response in the hemocoel of arthropods is essential for survival as it prevents the invasion of pathogens throughout the animal body. Antimicrobial peptides (AMPs) play an important role in this response by rapidly killing invading microorganisms. In this study, a novel cysteine-rich AMP has been isolated and characterized from the hemocytes of the cattle tick, Boophilus microplus. In addition to growth inhibition of Escherichia coli and Micrococcus luteus, the newly described AMP, designated ixodidin (derived from the Family Ixodidae), was found to exert proteolytic inhibitory activity against two exogenous serine proteinases, elastase and chymotrypsin. This is the first report of a molecule of an arachnid that has been shown to inhibit bacterial growth and proteinase activity.
Assuntos
Anti-Infecciosos/farmacologia , Peptídeos Catiônicos Antimicrobianos/farmacologia , Hemócitos/química , Ixodidae/química , Ixodidae/citologia , Peptídeos/farmacologia , Inibidores de Serina Proteinase/farmacologia , Sequência de Aminoácidos , Animais , Anti-Infecciosos/química , Anti-Infecciosos/isolamento & purificação , Peptídeos Catiônicos Antimicrobianos/química , Peptídeos Catiônicos Antimicrobianos/isolamento & purificação , Bovinos , Dados de Sequência Molecular , Peptídeos/química , Peptídeos/isolamento & purificação , Homologia de Sequência de Aminoácidos , Inibidores de Serina Proteinase/química , Inibidores de Serina Proteinase/isolamento & purificaçãoRESUMO
Lectins/carbohydrate binding can be involved in the Schistosoma mansoni recognition and activation of the Biomphalaria hemocytes. Therefore, expression of lectin ligands on Biomphalaria hemocytes would be associated with snail resistance against S. mansoni infection. To test this hypothesis, circulating hemocytes were isolated from B. glabrata BH (snail strain highy susceptible to S. mansoni), B. tenagophila Cabo Frio (moderate susceptibility), and B. tenagophila Taim (completely resistant strains), labelled with FITC conjugated lectins (ConA, PNA, SBA, and WGA) and analyzed under fluorescence microscopy. The results demonstrated that although lectin-labelled hemocytes were detected in hemolymph of all snail species tested, circulating hemocytes from both strains of B. tenagophila showed a larger number of lectin-labelled cells than B. glabrata. Moreover, most of circulating hemocytes of B. tenagophila were intensively labelled by lectins PNA-FITC and WGA-FITC, while in B. glabrata small hemocytes were labeled mainly by ConA. Upon S. mansoni infection, lectin-labelled hemocytes almost disappeared from the hemolymph of Taim and accumulated in B. glabrata BH. The role of lectins/carbohydrate binding in resistance of B. tengophila infection to S. mansoni is still not fully understood, but the data suggest that there may be a correlation to its presence with susceptibility or resistance to the parasite.
Assuntos
Biomphalaria/parasitologia , Hemócitos/química , Lectinas/metabolismo , Schistosoma mansoni/fisiologia , Animais , Biomphalaria/classificação , Contagem de Células , Interações Hospedeiro-Parasita , Microscopia de Fluorescência , FagocitoseRESUMO
Serine proteinases are involved, besides digestive role, in immune response processes. In addition to the typical serine proteinase domain, proteinases from arthropod haemocytes contain so-called clip domains which are believed to exert regulatory functions. Clones coding for clip domain-containing serine proteinases were isolated from both Penaeus vannamei and Penaeus monodon haemocyte cDNA libraries. These proteins have most of the structural characteristics of serine proteinase domain, but in the clip domain there are only four cysteines, whereas in most other clip domains there are six. Such structures are named pseudo-clip domains and apparently seem to be widely distributed in Penaeid shrimp. These proteinases were only expressed in haemocytes and not in muscles, hypodermis, heart, tail stalk, pleopods or hepatopancreas.
Assuntos
Hemócitos/química , Penaeidae/metabolismo , Serina Endopeptidases/sangue , Sequência de Aminoácidos , Animais , Sequência de Bases , Primers do DNA , Perfilação da Expressão Gênica , Biblioteca Gênica , Dados de Sequência Molecular , Penaeidae/genética , Estrutura Terciária de Proteína , Alinhamento de Sequência , Análise de Sequência de DNA , Serina Endopeptidases/genética , Serina Endopeptidases/metabolismo , Especificidade da EspécieRESUMO
Antimicrobial peptides (AMPs) are components of the immune system of both vertebrate and invertebrate animals. This study describes the isolation, primary structure, cDNA cloning, and tissue expression profile of two cysteine-rich AMPs from the hemolymph of the cattle tick Boophilus microplus. A 10,204 Da polypeptide, with six cysteine residues and no sequence similarity to any known molecule, was isolated from the cell-free hemolymph. Because of its sequence originality, this peptide was named microplusin. The second AMP was isolated from the hemocytes of B. microplus. This peptide, with a molecular mass of 4285 Da and six cysteines, is a defensin with similarity to the insect defensin family members. The cDNA cloning established that microplusin is synthesized as a prepeptide while the tick defensin is synthesized as a prepromolecule. Interestingly, despite the fact that microplusin and defensin have been isolated from different compartments, their gene expression was found to have similar tissue distribution.
Assuntos
Peptídeos Catiônicos Antimicrobianos/genética , Ixodidae/genética , Sequência de Aminoácidos , Animais , Peptídeos Catiônicos Antimicrobianos/isolamento & purificação , Peptídeos Catiônicos Antimicrobianos/farmacologia , Bovinos/parasitologia , Cromatografia Líquida de Alta Pressão , DNA Complementar/química , DNA Complementar/genética , Corpo Adiposo/química , Feminino , Expressão Gênica , Hemócitos/química , Hemolinfa/química , Testes de Sensibilidade Microbiana , Micrococcus luteus/efeitos dos fármacos , Dados de Sequência Molecular , Ovário/química , Reação em Cadeia da Polimerase , Análise de Sequência de DNA , Homologia de Sequência de AminoácidosRESUMO
We have purified a small size antimicrobial peptide, named gomesin, from the hemocytes of the unchallenged tarantula spider Acanthoscurria gomesiana. Gomesin has a molecular mass of 2270.4 Da, with 18 amino acids, including a pyroglutamic acid as the N terminus, a C-terminal arginine alpha-amide, and four cysteine residues forming two disulfide bridges. This peptide shows marked sequence similarities to antimicrobial peptides from other arthropods such as tachyplesin and polyphemusin from horseshoe crabs and androctonin from scorpions. Interestingly, it also shows sequence similarities to protegrins, antimicrobial peptides from porcine leukocytes. Gomesin strongly affects bacterial growth, as well as the development of filamentous fungi and yeast. In addition, we showed that gomesin affects the viability of the parasite Leishmania amazonensis.