Purification and Properties of Laticeptin, an Antimicrobial Peptide from Skin Secretions of the South American Frog Leptodactylus laticeps

Conlon, J. Michael; Al-Ghaferi, Nadia; Abraham, Bency; Sonnevend, Ágnes; King, Jay D.; Nielsen, Per F.

doi:10.2174/092986606775974410

Cited by 20 publications

(18 citation statements)

References 21 publications

(30 reference statements)

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“…Similarly to the observed for ocellatin-LB1 and -LB2, other AMPs such as leptoglycin and ocellatin-L1 (previously known as laticeptin), which were also isolated from the skin secretion of anurans, present restrict profiles of activities and are efficient only against gram-negative bacteria [21, 33]. According to Rollins-Smith et al [24], the presence of high amounts of peptides in the anuran skin secretion may compensate for their relatively low antimicrobial activities, since the peptide concentration may exceed the MIC values for many pathogens to which the animal is exposed in the wild.…”

Section: Resultsmentioning

confidence: 94%

Ocellatin peptides from the skin secretion of the South American frog Leptodactylus labyrinthicus (Leptodactylidae): characterization, antimicrobial activities and membrane interactions

Gusmao

Santos

et al. 2017

J Venom Anim Toxins Incl Trop Dis

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BackgroundThe availability of antimicrobial peptides from several different natural sources has opened an avenue for the discovery of new biologically active molecules. To the best of our knowledge, only two peptides isolated from the frog Leptodactylus labyrinthicus, namely pentadactylin and ocellatin-F1, have shown antimicrobial activities. Therefore, in order to explore the antimicrobial potential of this species, we have investigated the biological activities and membrane interactions of three peptides isolated from the anuran skin secretion.MethodsThree peptide primary structures were determined by automated Edman degradation. These sequences were prepared by solid-phase synthesis and submitted to activity assays against gram-positive and gram-negative bacteria and against two fungal strains. The hemolytic properties of the peptides were also investigated in assays with rabbit blood erythrocytes. The conformational preferences of the peptides and their membrane interactions have been investigated by circular dichroism spectroscopy and liposome dye release assays.ResultsThe amino acid compositions of three ocellatins were determined and the sequences exhibit 100% homology for the first 22 residues (ocellatin-LB1 sequence). Ocellatin-LB2 carries an extra Asn residue and ocellatin-F1 extra Asn-Lys-Leu residues at C-terminus. Ocellatin-F1 presents a stronger antibiotic potential and a broader spectrum of activities compared to the other peptides. The membrane interactions and pore formation capacities of the peptides correlate directly with their antimicrobial activities, i.e., ocellatin-F1 > ocellatin-LB1 > ocellatin-LB2. All peptides acquire high helical contents in membrane environments. However, ocellatin-F1 shows in average stronger helical propensities.ConclusionsThe obtained results indicate that the three extra amino acid residues at the ocellatin-F1 C-terminus play an important role in promoting stronger peptide-membrane interactions and antimicrobial properties. The extra Asn-23 residue present in ocellatin-LB2 sequence seems to decrease its antimicrobial potential and the strength of the peptide-membrane interactions.Electronic supplementary materialThe online version of this article (doi:10.1186/s40409-017-0094-y) contains supplementary material, which is available to authorized users.

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Section: Resultsmentioning

confidence: 94%

Ocellatin peptides from the skin secretion of the South American frog Leptodactylus labyrinthicus (Leptodactylidae): characterization, antimicrobial activities and membrane interactions

Gusmao

Santos

et al. 2017

J Venom Anim Toxins Incl Trop Dis

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“…After eliminating the tissue debris by centrifugation, the protein extracts were washed, and the aqueous phase containing the soluble proteins was used for the analysis. Protein samples were mixed with sample buffer, boiled for 3 min, and loaded on the well with the running buffer as described [34] using Tricine-SDS-PAGE containing separating gel (16 %), spacer gel (10 %), and stacking gel (4 %). Tricine-SDS-PAGE was the suitable method to separate peptide or protein with small mass.…”

Section: Protein Electrophoresis From Transformantsmentioning

confidence: 99%

Production of antibacterial peptide from bee venom via a new strategy for heterologous expression

et al. 2014

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Honey bee is important economic insect that not only pollinates fruits and crops but also provides products with various physiological activities. Bee venom is a functional agent that is widely applied in clinical treatment and pharmacy. Secapin is one of these agents that have a significant role in therapy. The functions of secapin from the bee venom have been documented, but little information is known about its heterologous expression under natural condition. Moreover, few scholars verified experimentally the functions of secapin from bee venom in vitro. In this study, we successfully constructed a heterologous expression vector, which is different from conventional expression system. A transgenic approach was established for transformation of secapin gene from the venom of Apis mellifera carnica (Ac-sec) into the edible fungi, Coprinus cinereus. Ac-sec was encoded by a 234 bp nucleotide that contained a signal peptide domain and two potential phosphorylation sites. The sequence exhibited highly homology with various secapins characterized from honey bee and related species. Southern blot data indicated that Ac-sec was present as single or multiple copy loci in the C. cinereus genome. By co-transformation and double-layer active assay, Ac-sec was expressed successfully in C. cinereus and the antibacterial activity of the recombinants was identified, showing notable antibacterial activities on different bacteria. Although Ac-sec is from the venom of Apidae, phylogenetic analysis demonstrated that Ac-sec was more closely related to that of Vespid than to bee species from Apidae. The molecular characteristics of Ac-sec and the potential roles of small peptides in biology were discussed.

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“…The search for similar sequences was performed with FASTA 3 program (http://www.ebi.ac.uk/fasta33/) using Swiss-Prot databank. New ocellatins were also off-line compared with fallaxin, pentadactylin, laticeptin and other ocellatins using information available in the current literature Rollins-Smith et al, 2005;Conlon et al, 2006), but unavailable in major databanks. Clustal W tool (http://www.ebi.ac.uk/clustalw/) was used to calculate the identity and similarity scores among the peptides.…”

Section: Sequence Analysismentioning

confidence: 99%

“…One of the most diverse and abundant frogs in the Neotropics is that of the genus Leptodactylus Fitzinger, 1826, which is divided into four species groups (Heyer and Reid, 2003). Despite the great diversity and wide distribution of this genus, studies on the skin secretion are rare, studied up to now: fallaxin (Leptodactylus fallax- Rollins-Smith et al, 2005), laticeptin (L. laticeps- Conlon et al, 2006), ocellatins (L. ocellatus- Nascimento et al, 2004Nascimento et al, , 2007, pentadactylin (L. pentadactylus- King et al, 2005) and syphaxins (L. syphax- Dourado et al, 2007). L. laticeps, L. ocellatus, L. pentadactylus and L. syphax are South American species, whereas L. fallax is from Central America.…”

mentioning

confidence: 99%

“…The molecular heterogeneity of ocellatins may be exploited as an aid to taxonomic and phylogenetic classification and in the design of peptide-based antimicrobial agents. The molecular diversity in the present-day species is a consequence of multiple duplications of this ancestral gene during radiation of the species and within individual species (Conlon et al, 2006).…”

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confidence: 99%

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Leptodactylus ocellatus (Amphibia): mechanism of defense in the skin and molecular phylogenetic relationships

et al. 2009

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Amphibian antimicrobial peptides have been known for many decades and several of them have already been isolated. However, the number of species investigated is still small. Herein, we report on the skin secretions of Leptodactylus ocellatus, which were extracted by mild electrical stimulation and its semi-preparative reverse-phase chromatography was resolved in more than 30 fractions. Among these fractions, two novel antimicrobial peptides were isolated and their amino acid sequences determined by de novo sequencing. The ocellatins-5 and -6 (21 and 22 amino acid residues, respectively) are amidated at the C-terminus. Ocellatins inhibited the growth of reference strains of both Gram-negative bacteria (Escherichia coli) and Gram-positive bacteria (Staphylococcus aureus) with minimal inhibition concentration values in the range of 32-128 microg/mL. The amino acid sequence of the peptides shows structural similarity with members of the antimicrobial peptides found in the skin secretion of other leptodactylid frogs. This observation is consistent with the hypothesis that many frog skin antimicrobial peptides are related evolutionarily, having arisen from multiple duplications of an ancestral gene that existed before the radiation of the different species.

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Purification and Properties of Laticeptin, an Antimicrobial Peptide from Skin Secretions of the South American Frog Leptodactylus laticeps

Cited by 20 publications

References 21 publications

Ocellatin peptides from the skin secretion of the South American frog Leptodactylus labyrinthicus (Leptodactylidae): characterization, antimicrobial activities and membrane interactions

Ocellatin peptides from the skin secretion of the South American frog Leptodactylus labyrinthicus (Leptodactylidae): characterization, antimicrobial activities and membrane interactions

Production of antibacterial peptide from bee venom via a new strategy for heterologous expression

Leptodactylus ocellatus (Amphibia): mechanism of defense in the skin and molecular phylogenetic relationships

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