Evaluating the Bioactivity of a Novel Broad-Spectrum Antimicrobial Peptide Brevinin-1GHa from the Frog Skin Secretion of Hylarana guentheri and Its Analogues

Abstract

Many antimicrobial peptides (AMPs) have been identified from the skin secretion of the frog Hylarana guentheri (H.guentheri), including Temporin, Brevinin-1, and Brevinin-2. In this study, an antimicrobial peptide named Brevinin-1GHa was identified for the first time by using ‘shotgun’ cloning. The primary structure was also confirmed through mass spectral analysis of the skin secretion purified by reversed-phase high-performance liquid chromatography (RP-HPLC). There was a Rana-box (CKISKKC) in the C-terminal of Brevinin-1GHa, which formed an intra-disulfide bridge. To detect the significance of Rana-box and reduce the hemolytic activity, we chemically synthesized Brevinin-1GHb (without Rana-box) and Brevinin-1GHc (Rana-box in central position). Brevinin-1GHa exhibited a strong and broad-spectrum antimicrobial activity against seven microorganisms, while Brevinin-1GHb only inhibited the growth of Staphylococcus aureus (S. aureus), which indicates Rana-box was necessary for the antimicrobial activity of Brevinin-1GHa. The results of Brevinin-1GHc suggested transferring Rana-box to the central position could reduce the hemolytic activity, but the antimicrobial activity also declined. Additionally, Brevinin-1GHa demonstrated the capability of permeating cell membrane and eliminating biofilm of S. aureus, Escherichia coli (E. coli), and Candida albicans (C. albicans). The discovery of this research may provide some novel insights into natural antimicrobial drug design.

Brevinin-1GHd: a novel Hylarana guentheri skin secretion-derived Brevinin-1 type peptide with antimicrobial and anticancer therapeutic potential

“…As the NCBI-BLAST alignment suggested, this novel peptide shared similar properties with most peptides in the Brevinin-1 subfamily. Among these, Brevinin-1GHd is 83% identical with Brevinin-1GHa and 71% identical with Gaegurin-5 [13,14].…”

“…In our previous study, one novel Brevinin-1 type peptide, Brevinin-1GHa, was first found in Hylarana guentheri and two analogs, Brevinin-1GHb and Brevinin-1GHc, were designed to further study the relationships between structure and activity. The study of Brevinin-1GHa and its two analogs revealed that the deletion or removal of the "Rana-Box" decreased its antimicrobial activity while reducing hemolytic activity [13]. Through comparing differences of structure and activity between Brevinin-1GHa and its analogs, the hypothesis was established that the hemolytic activity was determined both by the hydrophobicity and hydrophobic face since the lack and decrease in hydrophobicity of Brevinin-1GHb and Brevinin-1GHc showed lower hemolytic activity than Brevinin-1GHa [13].…”

“…The study of Brevinin-1GHa and its two analogs revealed that the deletion or removal of the "Rana-Box" decreased its antimicrobial activity while reducing hemolytic activity [13]. Through comparing differences of structure and activity between Brevinin-1GHa and its analogs, the hypothesis was established that the hemolytic activity was determined both by the hydrophobicity and hydrophobic face since the lack and decrease in hydrophobicity of Brevinin-1GHb and Brevinin-1GHc showed lower hemolytic activity than Brevinin-1GHa [13]. Interestingly, although Brevinin-1GHd had the same hydrophobic face, higher hydrophobicity and similar length of alpha-helix structure as Brevinin-1GHa, its hemolytic activity was slightly lower than Brevinin-1GHa ( Figure 8).…”

“…In a previous study, we also characterized and designed Brevinin-1GHa and its two analogs to further study the relationship of the "Rana-Box" structure and its effect on antimicrobial activity. However, neither truncating the C-terminal cyclic structure nor removing the "Rana-Box" to the middle part of the peptide, greatly decreased antimicrobial activity [13]. The actual role of the "Rana-Box" in antimicrobial performance is still ambiguous and more factors need to be considered.…”

Bv8-Like Toxin from the Frog Venom of Amolops jingdongensis Promotes Wound Healing via the Interleukin-1 Signaling Pathway

“…This characteristic makes them to have extreme chemical diversity [7,8]. Till now, several biochemical arsenals with potential pharmaceutical properties, particularly antimicrobial peptides (AMPs), have been discovered from the skin secretions of amphibians [9][10][11][12][13][14]. Despite this, little information is available about amphibian skin-derived bioactive compounds with wound healing activity.…”