Modification Targeting the “Rana Box” Motif of a Novel Nigrocin Peptide From Hylarana latouchii Enhances and Broadens Its Potency Against Multiple Bacteria

Abstract

Public health is confronting the threat caused by antibiotic resistance and this means new antibacterial strategies must be developed urgently. Antimicrobial peptides (AMPs) have been considered as promising therapeutic candidates against infection in the post-antibiotic era. In this paper, we dismissed the significance of “Rana box” in the natural nigrocin-HL identified from skin secretion of Hylarana latouchii by comparing its activity with nigrocin-HLD without the motif. By substituting the “Rana box” sequence with an amidated phenylalanine residue, the natural peptide was modified into a shorter AMP nigrocin-HLM. Activities and toxicities of these two peptides in vitro and in vivo were compared. As a result, nigrocin-HLM not only displayed significantly increased potency against several representative microbes, but also high activity against the antibiotic-resistant methicillin-resistant S. aureus (MRSA, NCTC 12493 and ATCC43300 and several clinical isolates) as evidenced by markedly reduced minimal inhibitory concentration (MIC), minimal bactericidal concentration (MBC), and minimum biofilm eradication concentration (MBEC). More strikingly, nigrocin-HLM exhibited prominent inhibition against MRSA infection in a pneumonia mice model. In addition, the substitution attenuated the toxicity of nigrocin-HLM as evidenced by precipitously decreased hemolytic and cytotoxic activities in vitro, and acute toxicity to mice in vivo. Taken these results into consideration, nigrocin-HLM should be a promising therapeutic candidate for anti-infection. And in addition to dismiss an indispensable role of “Rana box” in maintaining antimicrobial activity of nigrocin-HL, our data provided an inspired strategy for peptide optimization.

Antimicrobial Peptides: Classification, Design, Application and Research Progress in Multiple Fields

“…The deletion of the 'Rana box' motif will cause the AMP antibacterial effect to disappear, but replacing the natural 'Rana box' sequence of AMPs with amidated phenylalanine can expand its efficacy against antibiotic-resistant microorganisms, including MRSA and P. aeruginosa, and reduce cytotoxicity. This phenomenon also shows that the effect of the motif on AMPs needs to be determined based on the specific situation and is not completely beneficial (Bao et al, 2018).…”

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

“…In fact, no matter what changes in this cyclic structure were made, these would always come with some variations on structural parameters or secondary structure. For example, truncating the "Rana-Box" greatly increased the helicity of Nigrocin-HL analog, which may explain why the antimicrobial activity of Nigrocin-HLD was better [12]. However, the elimination of cyclic structure made the putative length of the alpha-helix of Brevinin-1GHb shorter than Brevinin-1GHa, which was in accordance with the decrease of antimicrobial activity.…”

“…Furthermore, the cytotoxicity of Brevinin-1GHd against human microvascular endothelial cells (HMEC-1) (ATCC-CRL-3243) and human keratinocytes (HaCaT) (ATCC-PCS-200-011) were evaluated as reported [12]. HMEC-1 cell line (5 × 10 3 cells/ml) was seeded into a 96-well plate with MCDB131 medium (Life technologies, Carlsbad, CA) supplemented with 10% (v/v) fetal bovine serum (FBS), 1% (v/v) penicillin-streptomycin, 10 ng/ml epidermal growth factor, 10 mM L-glutamine and 1 μg/ml hydrocortisone for 24 h. HaCaT (5 × 10 3 cells/ml) was seeded into a 96-well plate with DMEM medium (Life technologies, U.K.) supplemented with 10% (v/v) FBS, 1% (v/v) penicillin-streptomycin.…”

Identification and Target-Modification of SL-BBI: A Novel Bowman–Birk Type Trypsin Inhibitor from Sylvirana latouchii

“…The active substances in amphibian skin secretions have always occupied an essential position in the field of natural drug discovery. Currently, a variety of bioactive peptides of brevinin, temporin, ranatensin, nigrocin and ranacyclin families have been discovered from broad-folded frog (Sylvirana latouchii) [31][32][33][34] and showed potential medicinal value. Here, we found a novel Bowman-Birk type trypsin inhibitor peptide from this species named SL-BBI, which shared the highly conserved '-CWTP 1 SXXPKPC-' ring with the ranacyclin family.…”