Abstract
Liver-expressed antimicrobial peptides (LEAPs) are cysteine-containing cationic peptides. LEAP-1 and LEAP-2 are eight- and four-cysteine containing antimicrobial peptides found in animals, respectively. LEAP-1 is widely known as antibacterial peptide involved in the innate immunity of fish, but the roles of LEAP-1 and LEAP-2 in Antarctic fish species are unknown. In the present study, we synthesized and characterized novel LEAPs with four and eight cysteine residues, derived from Antarctic notothenioid (Dissostichus mawsoni) and Antarctic eelpout (Lycodichthys dearborni). Circular dichroism spectroscopy of these peptides showed a typical β-sheet conformation. The LEAPs were found to be bactericidal against gram-positive as well as gram-negative bacteria. In the SYTOX green uptake assay, LEAPs did not trigger any significant increase in fluorescence. However, LEAPs competitively bound to DNA and replaced the ethidium bromide (EB) dye. To determine the effect of temperature on the activity of LEAPs, we evaluated the antibacterial activity against Listeria monocytogenes at 5, 15, 25, and 35 °C. The results showed that the antibacterial activity of LEAPs increased with a decrease in temperature, which may indicate that the Antarctic fish LEAP are evolutionarily adapted. Taken together, our results suggest that novel Antarctic LEAPs are bactericidal peptides with the likely mode of action being DNA binding and may be evolved to adapt to cold temperature.
Highlights
Liver-expressed antimicrobial peptides (LEAPs) are cysteine-containing cationic peptides with antimicrobial activity, which were first discovered in human blood ultrafiltrate and urine samples [1].In humans, LEAPs with eight and four cysteine residues are termed as hepcidin [1] or LEAP-1 [2]and LEAP-2 [3], respectively
The amino terminal Cu(II)and Ni(II)-binding (ATCUN) motif (QSH) was found to be present in nLEAP-1, which could lead to increased interaction with DNA, and a more potent antibacterial activity
Incubation of the nLEAP-2 and eLEAP-2 at different temperatures from 5 to 35 ◦ C showed a change in the Circular Dichroism (CD) spectrum at low temperatures, indicating a modification of the peptide structure
Summary
Liver-expressed antimicrobial peptides (LEAPs) are cysteine-containing cationic peptides with antimicrobial activity, which were first discovered in human blood ultrafiltrate and urine samples [1].In humans, LEAPs with eight and four cysteine residues are termed as hepcidin [1] or LEAP-1 [2]and LEAP-2 [3], respectively. The human LEAP-1 (hLEAP-1) is a cysteine-containing cationic peptide comprising about 20–25 amino acid residues [2]. It has a characteristic β-sheet structure, which is stabilized by eight cysteine residues [2]. These residues form four disulfide bonds, one of which is an unusual vicinal bond between adjacent cysteines at the hairpin turn with an amino terminal Cu(II)and Ni(II)-binding (ATCUN) motif [4]. The N-terminal region of human LEAP-1 (hepcidin-25) has a metal binding site specific for the coordination of Cu(II) and Ni(II) known as ATCUN motif [5]
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