Abstract
Recently, defense peptides that are able to act against several targets have been characterized. The present work focuses on structural and functional evaluation of the peptide analogue Pa-MAP, previously isolated as an antifreeze peptide from Pleuronectes americanus. Pa-MAP showed activities against different targets such as tumoral cells in culture (CACO-2, MCF-7 and HCT-116), bacteria (Escherichia coli ATCC 8739 and Staphylococcus aureus ATCC 25923), viruses (HSV-1 and HSV-2) and fungi (Candida parapsilosis ATCC 22019, Trichophyton mentagrophytes (28d&E) and T. rubrum (327)). This peptide did not show toxicity against mammalian cells such as erythrocytes, Vero and RAW 264.7 cells. Molecular mechanism of action was related to hydrophobic residues, since only the terminal amino group is charged at pH 7 as confirmed by potentiometric titration. In order to shed some light on its structure-function relations, in vitro and in silico assays were carried out using circular dichroism and molecular dynamics. Furthermore, Pa-MAP showed partial unfolding of the peptide changes in a wide pH (3 to 11) and temperature (25 to 95°C) ranges, although it might not reach complete unfolding at 95°C, suggesting a high conformational stability. This peptide also showed a conformational transition with a partial α-helical fold in water and a full α-helical core in SDS and TFE environments. These results were corroborated by spectral data measured at 222 nm and by 50 ns dynamic simulation. In conclusion, data reported here show that Pa-MAP is a potential candidate for drug design against pathogenic microorganisms due to its structural stability and wide activity against a range of targets.
Highlights
Advances in molecular biology have explained the complexity of life while retaining the simplicity of biology’s central dogma
Pa-multiple active peptides (MAPs) was purified by semi-preparative reversed-phase chromatography with linear acetonitrile gradient of 5 to 95% ACN over water, with 0.1% trifluoroacetic acid (TFA) (Figure 1A)
The understanding of Pa-MAP structural stability and conformational preference at the molecular level in a hydrophobic environment may lead to advances in drug design and therapy
Summary
Advances in molecular biology have explained the complexity of life while retaining the simplicity of biology’s central dogma. A comparison between AFPs and antimicrobial peptides (AMPs) has revealed similar structural and physical-chemical properties, including hydrophobic ratio (45 to 55% hydrophobicity), polypeptide chain length (twenty to thirty amino acid residues), hydrophobic moment and a specific amino acid composition (mainly threonine, leucine, histidine, lysine, arginine, aspartic acid and alanine) [7]. These characteristics probably reflect the fact that many of these peptides may be capable of interacting with and disrupting target cell membranes [8]. Considering these shared characteristics, the key question is: Do these fish-sourced peptides have a single function? It seems that they do not, since the concept of peptides with multiple functions has been commonly observed in different organisms, representing savings in energy for organisms that express a single gene with multiple activities [1,9]
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