ANTIMICROBIAL PEPTIDES FROM AMPHIBIAN SKIN: WHAT DO THEY TELL US?

Amphibian skin secretions contain many biologically active compounds, such as biogenic amines, complex alkaloids, or peptides. Within the latter class of molecules, a large number of peptide antibiotics has been isolated and characterized from different amphibian species. Antimicrobial peptides are considered the effector molecules of innate immunity, acting as a first line of defense against bacterial infections, by perturbing the phospholipid bilayer of the target cell membrane. These gene-encoded molecules are synthesized as inactive precursors and in several cases their proparts were shown to have highly conserved structures. It has also been demonstrated that the promoter regions of inducible peptide antibiotics are often regulated by the transcriptional control machinery NF-kB/IkBa. In amphibia of Rana and Bombina genera, inhibition of transcription of the genes encoding antimicrobial peptides has been obtained by glucocorticoid treatment, which causes an increase of IkBa synthesis. Moreover, determination of the structure of a number of genes coding for antimicrobial peptides in amphibia has actually shown that their promoter regions contain recognition sites for nuclear factors

An amphibian antimicrobial peptide variant expressed in Nicotiana tabacum confers resistance to phytopathogens.

Esculentin-1 is a 46-residue antimicrobial peptide present in skin secretions of Rana esculenta. It is effective against a wide variety of micro-organisms, including plant pathogens with negligible effects on eukaryotic cells. As a possible approach to enhance plant resistance, a DNA coding for esculentin-1, with the substitution Met-28Leu, was fused at the C-terminal end of the leader sequence of endopolygalacturonase-inhibiting protein, under the control of the cauliflower mosaic virus 35S promoter region, and introduced into Nicotiana tabacum. The antimicrobial peptide was isolated from the intercellular fluids of healthy leaves of transgenic plants, suggesting that it was properly processed, secreted outside cells and accumulated in the intercellular spaces. The morphology of transgenic plants was unaffected. Challenging these plants with bacterial or fungal phytopathogens demonstrated enhanced resistance up to the second generation. Moreover, transgenic plants displayed insecticidal properties

Evoluzione dei carotenoidi nel corso della maturazione di uve Tintilia

I carotenoidi presenti nelle uve, principalmente luteina e \uf062-carotene, sono importanti precursori di aromi caratteristici, appartenenti alla classe dei C13-norisoprenoidi. Tali composti sono di grande interesse per la loro bassa soglia olfattiva e perch\ue9 contribuiscono in modo significativo alla qualit\ue0 e tipicit\ue0 dei vini. Nella presente nota si \ue8 valutata l\u2019evoluzione dei carotenoidi nel corso della maturazione di uve Tintilia, vitigno fortemente legato al territorio della regione Molise, in relazione alla forma di allevamento. La forma di allevamento a Guyot, rispetto al cordone speronato, ha favorito un maggior accumulo di sostanze polifenoliche, antocianiche e di carotenoidi nelle uve. La luteina ha registrato la pi\uf9 alta concentrazione, circa il 60% dei carotenoidi totali, con una riduzione di circa il 35% nei due mesi prima della raccolta

Expression and activity of cyclic and linear analogues of esculentin-1, an antimicrobial peptide from amphibian skin.

Esculentin-1 is a potent anti-microbial peptide present in minute amounts in skin secretions of Rana esculenta. It contains 46 amino-acid residues and a C-terminal disulfide bridge. We have explored the possibility of producing analogues of this peptide by recombinant expression in Escherichia coli of a fusion protein which is sequestered in inclusion bodies. The peptide of interest has been inserted at the N-terminus of the protein, from which it can be released by cyanogen bromide cleavage. The anti-microbial activities of the recombinant peptide as well as that of a mutant linear form devoid of the disulfide bridge are presented. The recombinant analogues retain the biological activity of the natural peptide, as tested with an inhibition zone assay against a variety of microorganisms. However, experiments on the rate of bacterial killing show that gram-negative bacteria are more sensitive to the peptides than the gram-positive bacterium, the effect of the cyclic peptide being in all cases faster than that of the linear molecule. Moreover, the activity against gram-negative bacteria for both peptides is not affected by salts, whereas the activity against Staphylococcus aureus is lost at high salt concentration

Surface Plasmon Resonance technology to assess biological interactions

Molecular interactions between proteins or between proteins and small molecules are pivotal events for selective binding of biological structures and, consequentially, for their correct function. In this scenario, the evaluation of kinetic parameters, characterizing a molecular interactions, is considered a crucial event to reveal the nature of binding processes. The focus on peculiar forces involved in the molecular recognition represents an opportunity to explore biological interactions in real time, and to develop a number of innovative biotechnological methods for diagnosis and/or therapy. Currently, optical biosensors, offering an increasingly effective technology to detect in real time molecular binding, are usually composed by a detector, a sensor surface and a sample delivery system: only definite substances, which are able to interact specifically with the biological part, lead to an optical or electrical signal of the physical transducer. In this review we want to highlight the exponentially-growing interest of Surface Plasmon Resonance (SPR) based optical biosensors for molecular binding analysis in different research fields

Tachykinins and other biologically active peptides from the skin of the Costa Rican phyllomedusid frog Agalychnis callidryas.

Peptides present in a methanol extract prepared from skin of the Costa Rican frog Agalychnis callidryas of the Phyllomedusinae subfamily were studied by sequence analysis and pharmacological tests. Members of five different peptide families-tachykinins, bradykinins, caerulein, opioid peptides and sauvagine-were found. In particular, the extract contained a number of tachykinins with the following sequences: Gly-Pro-Pro-Asp-Pro-Asn-Lys-Phe-Ile-Gly-Leu-Met-NH2, Gly-Pro-Pro-Asp-Pro-Asp-Arg(Lys)-Phe-Tyr-Pro-Gly-Met-NH2, pGlu-Pro-Asp-Pro-Asp-Arg-Phe-Tyr-Pro-Gly-Met-NH2, Gly-Pro-Pro-Asp-Pro-Asn-Lys-Phe-Tyr-Pro-Val-Met. The latter three peptides have the unusual C-terminal sequence Pro-Gly(or Val)-Met-NH2 rather than Gly-Leu-Met-NH2 found in many other members of the tachykinin family. The observed amino acid substitutions may be the reason for the marked decrease in the biological activity observed in all in vitro and in vivo tests, even through the spectrum of tachykinin activities was retained. A kassinin-like peptide, with the sequence Gly-Pro-Pro-Asp-Pro-Asn-Lys-Phe-Ile-Gly-Leu-Met-NH2, was also found in the A. callidryas skin. While kassinin has a much higher affinity for NK-3 than for NK-1 receptors, the opposite is true for this A. callidryas peptide. The extract from A. callidryas skin also contained a new caerulein (pGlu-Asp-Tyr(HSO3)-Lys-Gly-Trp-Met-Asp-Phe-NH2) and a phyllokinin (Arg-Pro-Hyp-Gly-Phe-Ser-Pro-Phe-Arg-Ile-Tyr), as well as the opioid peptides dermorphin and [Hyp6]dermorphin, both previously isolated from different Phyllomedusa species

Structure-function relationships in bombinins H, antimicrobial peptides from Bombina skin secretions.

Skin secretions of amphibia of the Bombina genus contain two families of antimicrobial peptides, the bombinins (bombinin-like peptides)and the bombinins H (H for hydrophobic and hemolytic). The latter family includes a number of peptides containing a D-amino acid in the second position, in addition to their corresponding all L-isomers. The antimicrobial activity of three pairs of bombinin H isomers, H2/H4,H6/H7 and GH-1D/GH-1L, has been investigated. The first two pairs of peptides were actually isolated from the secretion, whereas the third was synthesized according to the sequence deduced from a gene coding for a bombinin-like peptide in Bombina orientalis