Role of protein present in the ink released by-the-sea slug Aplysia dactylomela Rang in 1828 in defense mechanisms of the animal / Papel de proteínas presentes na tinta liberada pela lesma-do-mar Aplysia dactylomela Rang 1828 nos mecanismos de defesa do animal

AUTOR(ES)
DATA DE PUBLICAÇÃO

2005

RESUMO

The marine gastropod Aplysia dactylomela is known to release a purple ink when it is plagued. How has no external structure of protection, it is believed that secretion, rich in biologically active substances, participate in the chemical defense of the animal. The ink is composed of pigments, proteins and low molecular weight substances. The pigments of ink are originated from red algae, but as the protein, little is known about its origin, processing, storage location and function in animals. This paper describes for the first time the protein composition of the ink and presents some physico-chemical and dactylomelina of organic-P, an antibacterial protein present in the ink that slug, and get information about the location of this protein in the ink gland. The paint was obtained from specimens found on the beach in Fleixeiras, Ce. The protein composition of the ink was determined by two-dimensional electrophoresis and purification of dactylomelina-P was done by ion exchange chromatography and hydrophobic interaction. Dactylomelina-P was analyzed on the molecular mass, isoelectric point, composition of amino acids, the presence of carbohydrates, thermal stability, resistance to proteases and pH. Several biological activities were tested with the ink and the dactylomelina-P, including antimicrobial activity, enzyme, haemagglutinating, anticoagulant, hemolytic, cytotoxic and toxic. Tests for localization of the protein were performed in different tissues of the slug by western blot in the ink gland by immunohistochemistry and the interaction with the bacterium Staphylococcus was performed by immunocytochemistry. The ink of A. dactylomela contains more than 40 proteins / peptides, with masses below 70 kDa and PIs in the acidic range. The most abundant protein in the ink is dactylomelina-P, which is a monomeric molecule of 59.8 kDa, pI 5.0, which has high content of methionine and less than 1% of carbohydrates, is denatured at 60 C for 10 minutes and resist the pH between 3-12. Dactylomelina-P shows a broad spectrum of antibacterial action, but not antifungal, unlike paint that has a factor with this activity. It is particularly effective against marine bacteria and may be bactericidal (4.0 g / ml) or bacteriostatic (0.2 g / ml), depending on the concentration. P-Dactylomelina agglutinate rabbit erythrocytes, rats and mice, showed no anticoagulant activity, and cytotoxic and hemolytic. The LD50 for mice was between 60-100 mg / kg, is considered moderately toxic. Dactylomelina-P was found in the ink gland, is located preferentially in cells producing the vesicles. Tests for transmission electron microscopy revealed that the dactylomelina through the cell wall of the bacterium S. aureus and mainly interacts with the cytoplasmic membrane, probably interfering with metabolism, rather than damage the cell.

ASSUNTO(S)

dactylomelina-p - protein gastropods, aplysia dactylomela aplysia dactylomela microbiologia gastrópode marinho dactylomelina-p - proteína

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