Modified versions of an antimicrobial peptide from climbing perch fish showed stronger antibacterial activity against Gram-positive bacteria while remaining safe for human cells at therapeutic concentrations.
Safe at 20 µg/mLBoth engineered peptide variants showed enhanced antibacterial activity while remaining non-toxic to human fibroblast and Vero cell lines at therapeutic concentrations
What the researchers found
Two engineered variants of the AtMP2 peptide (AtMP2-1 and AtMP2-2), generated through systematic directed evolution, demonstrated higher antimicrobial activity against Gram-positive bacteria than the parent AtMP2 peptide, as measured by Minimum Inhibitory Concentration (MIC) and Kirby-Bauer disk diffusion assays. Activity against Gram-negative bacteria was comparatively lower.
Cytotoxicity testing using SRB assays on HS-27 (human fibroblast) and Vero cell lines showed both variants were safe at 20 µg/mL. Molecular docking analysis revealed strong binding interactions with bacterial proteins involved in cell death pathways, including SecA, RpoB, GyrA, ClpP, and MetG, with more negative scores indicating stronger binding.
Why it matters
Antimicrobial resistance is a global health crisis, and antimicrobial peptides (AMPs) represent one of the most promising alternatives to conventional antibiotics. Fish-derived AMPs are particularly interesting because fish constantly live in bacteria-rich aquatic environments and have evolved potent natural defenses. Showing that directed evolution can enhance these natural peptides' antibacterial potency while maintaining safety opens a pathway toward developing new anti-infective agents.
How the study worked
Researchers extracted antimicrobial peptides from the epidermal mucus of climbing perch (Anabas testudineus). The AtMP2 peptide was selected and subjected to systematic directed evolution to generate variants. Characterization was performed using computational tools (APD3, CAMP, AMPFun). Two promising variants (AtMP2-1 and AtMP2-2) were synthesized and tested for antibacterial activity via MIC determination and Kirby-Bauer disk diffusion against Gram-positive and Gram-negative bacteria. Cytotoxicity was assessed using SRB assays on HS-27 and Vero cell lines. Molecular docking (ZDOCK and HPEPDOCK) evaluated binding to bacterial target proteins.
What this study cannot tell us
This is an in vitro study — no animal infection models were used to test efficacy in a living system. The enhanced activity was mainly against Gram-positive bacteria, with limited improvement against Gram-negative organisms (which are often the more challenging clinical targets). Specific MIC values were not provided in the abstract. The molecular docking results are computational predictions that require experimental validation. Stability, pharmacokinetics, and in vivo toxicity remain untested.
How to read the evidence
This is an early-stage in vitro and computational study. While the directed evolution approach is well-established and the antibacterial testing methods are standard, the lack of in vivo data, unspecified MIC values, and reliance on computational docking for mechanism of action limit the evidence strength.
When this study was published
Published in 2025, this study is very recent and reflects current trends in antimicrobial peptide engineering. The use of directed evolution and computational characterization represents modern approaches to AMP development.
The bigger picture
The antimicrobial peptide field is actively seeking ways to improve natural AMPs for clinical use. This study demonstrates that directed evolution — a powerful protein engineering approach — can successfully enhance fish-derived peptides. If these enhanced peptides can be developed further with in vivo testing, they could contribute to the growing arsenal of non-antibiotic antimicrobials needed to combat drug-resistant infections.
Questions still open
- Can further rounds of directed evolution improve activity against Gram-negative bacteria, which pose greater clinical challenges?
- How do these peptides perform in animal infection models with realistic dosing and delivery?
- Could these fish-derived AMPs be combined with conventional antibiotics for synergistic effects against resistant bacteria?
Common questions
Why are scientists looking at fish mucus for new antibiotics?
Could these fish peptides replace antibiotics?
Read the original research
Directed Evolution of AtMP2 Peptide: Unlocking Enhanced Antibacterial Potential from Anabas testudineus.
Molecules (Basel, Switzerland), 30(23)
Citation
Lee, Li Ting; Ang, Arnold; Najm, Ahmed; Adnan, Adura Mohd; Nordin, Akram Mohd; Mahmood, Ibrahim; Dunkhorol, Sarantuya; Fazry, Shazrul; Law, Douglas. (2025). Directed Evolution of AtMP2 Peptide: Unlocking Enhanced Antibacterial Potential from Anabas testudineus.. Molecules (Basel, Switzerland), 30(23). https://doi.org/10.3390/molecules30234590