Dermaseptin-like peptide

Analysis of the Schistosoma mansoni peptidome for immunomodulatory molecules by solvent extraction and reverse-phase HPLC revealed a 27-amino-acid residue peptide from an extract of cercariae. Using matrix-assisted, laser desorption-ionization, time-of-flight mass spectrometry, the peptide yielded a protonated molecular ion [M + H]+ of m/z 2789. The unequivocal sequence was deduced by automated Edman degradation as: DLWNSIKDMAAAAGRAALNAVTGMVNQ. The peptide exhibited an 80.76% identity with dermaseptin 3.1 from the leaf frog Agalychnis annae, and was therefore named Schistosoma mansoni dermaseptin-like peptide (SmDLP). Immunocytochemical staining using a primary antidermaseptin B2 antibody located SmDLP in acetabular glands of cercariae, in and around schistosomula, and in adult worms and their eggs. Dot-blotting confirmed its presence in extracts (cercariae and worms) and excretion/secretion (E/S) products (transforming cercariae and eggs). This was corroborated by use of a MALDI-ToF spectra database of E/S products from cercariae. Functional characterization of the peptide indicated that SmDLP had typical amphipathic antimicrobial peptide properties, i.e., the ability to lyse human erythrocytes causing a decrease in the levels of nitric oxide produced by monocytic cells. This last function strongly suggests that SmDLP plays a vital role in the parasite's immunoevasion strategy. The possibility that schistosomes acquired this gene from amphibians has been discussed by constructing a phylogenetic tree.

The dermaseptin antimicrobial peptide family contains members of 27-34 amino acids in length that have been predominantly isolated from the skins/skin secretions of phyllomedusine leaf frogs. By use of a degenerate primer in Rapid amplification of cDNA ends (RACE) PCR designed to a common conserved domain within the 5'-untranslated regions of previously-characterized dermaseptin encoding cDNAs, two novel members of this peptide family, named dermaseptin-PD-1 and dermaseptin-PD-2, were identified in the skin secretion of the phyllomedusine frog, Pachymedusa dacnicolor. The primary structures of both peptides were predicted from cloned cDNAs, as well as being confirmed by mass spectral analysis of crude skin secretion fractions resulted from reversed-phase high-performance liquid chromatography. Chemically-synthesized replicates of dermaseptin-PD-1 and dermaseptin-PD-2 were investigated for antimicrobial activity using standard model microorganisms (Gram-positive bacteria, Gram-negative bacteria and a yeast) and for cytotoxicity using mammalian red blood cells. The possibility of synergistic effects between the two peptides and their anti-cancer cell proliferation activities were assessed. The peptides exhibited moderate to high inhibition against the growth of the tested microorganisms and cancer cell lines with low haemolytic activity. Synergistic interaction between the two peptides in inhibiting the proliferation of Escherichia coli and human neuronal glioblastoma cell line, U251MG was also manifested.

High-resolution mass spectrometry-based peptidomics has been used to characterize several components in electro-stimulated skin secretions of the endemic Mexican frog Pachymedusa dacnicolor. Peptide mass screening performed in an Orbitrap-XL mass spectrometer showed that P. dacnicolor skin secretions possess 194 different components with molecular masses ranging mainly from 500 to 6,000 Da. Dozens of molecules were partially sequenced including two novel protease inhibitors. Additionally, one posttranslationally modified bradykinin and two novel dermaseptin-like antimicrobial peptides were fully sequenced. The novel peptide named here DMS-DA5 was fully characterized and showed potent antibacterial activity against various bacteria such as Escherichia coli, Bacillus subtilis, Salmonella enterica serovar typhimurium, and Pseudomonas aeruginosa with minimal inhibitory concentrations from 3.10 to 25.0 microM.