Abstract
The natural peptide chensinin-1 doesnot exhibit its desired biological properties. In this study, the mutant MC1-1 was designed by replacing Gly in the chensinin-1 sequence with Trp. Mutants MC1-2 and MC1-3 were designed based on the MC1-1 sequence to investigate the specific role of His residues. The mutated peptides presented α-helicity in a membrane-mimetic environment and exhibited broad-spectrum antimicrobial activities; in contrast to Trp residues, His residues were dispensable for interacting with the cell membrane. The interactions between the mutant peptides and lipopolysaccharide (LPS) facilitated the ingestion of peptides by Gram-negative bacteria. The binding affinities of the peptides were similar, at approximately 10 μM, but ΔH for MC1-2 was −7.3 kcal.mol−1, which was 6-9 folds higher than those of MC1-1 and MC1-3, probably due to the conformational changes. All mutant peptides demonstrated the ability to inhibit LPS-induced tumour-necrosis factor-α (TNF-α) and interleukin-6 (IL-6) release from murine RAW264.7 cells. In addition, the representative peptide MC1-1showed better inhibition of serum TNF-α and IL-6 levels compared to polymyxin B (PMB), a potent binder and neutralizer of LPS as positive control in LPS-challenged mice model. These data suggest that the mutant peptides could be promising molecules for development as chensinin-based therapeutic agents against sepsis.
Highlights
In our previous work, the 18 amino acid antimicrobial peptide chensinin-1 was purified from the skin secretions of the Chinese brown frog Rana chensinensis and characterized[19,20]
When the His residues were replaced by Arg residues, the mean hydrophobicity value increased slightly to 9.788 (MC1-3), and the net charge increased to +1 0
The well-known LPS antagonistPMB, employed as a control experiment, reduced the TNF-a and IL-6 levels by 70% and 71%, respectively. These results indicated that all the peptides showed the ability to inhibit the production of TNF-αand IL-6, and mean hydrophobicity value was increased to 11.626 (MC1-2) displayed greater inhibition than PMB
Summary
The 18 amino acid antimicrobial peptide chensinin-1 was purified from the skin secretions of the Chinese brown frog Rana chensinensis and characterized[19,20]. Arg residues can confer positive charge and can form hydrogen bonds as well as cation-πinteractions, which render the insertion of Arg residues into a lipid bilayer more favorable[23] These features unique to Trp and Arg residues make them attractive molecules for use in the design of short-chain antimicrobial peptides. Previous studies indicated that the introduction of Trp residues can significantly improve the antimicrobial activity of the peptide against the Gram-negative bacteria, such as the design of the peptide of IK-6 analogues and chensinin-1b10,24. To improve the antimicrobial and anti-inflammatory activities of chensinin-1 and to better understand the specific biology function of His residues in the sequence of chensinin-1, we replaced three Gly residues with Trp to improve the hydrophobicity of the peptide and obtained the novel mutant peptide MC1-1. The capability of the mutated peptides to block LPS-dependent TNF-αand IL-6 secretion by mouse RAW 264.7 macrophages in vitro, as well as mice displaying endotoxemia mice in vivo were investigated
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