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Evolution-Based Protein Engineering for Antifungal Peptide Improvement
Jing Gu1,2, Noriyoshi Isozumi3, Shouli Yuan1,2
1Group of Peptide Biology and Evolution, State Key Laboratory of Integrated Management of Pest Insects and Rodents, Institute of Zoology, Chinese Academy of Sciences, Beijing, China.
Molecular Biology and Evolution
|July 28, 2021
Summary
Researchers engineered antimicrobial peptides (AMPs) to combat fungal infections. An evolution-based strategy identified a key site (UEAMS) that significantly boosted antifungal activity, offering a promising alternative to conventional antibiotics.
Area of Science:
- Biochemistry
- Molecular Biology
- Drug Discovery
Background:
- Antimicrobial peptides (AMPs) are promising antibiotic alternatives due to low resistance.
- Clinical application of AMPs is limited by low activity, high cost, and cytotoxicity.
- Engineering AMPs is challenging due to complex sequence-structure-function relationships.
Purpose of the Study:
- To develop an evolution-based strategy for enhancing antifungal activity of AMPs.
- To identify and characterize sites responsible for improved antifungal efficacy.
- To engineer a nematode-sourced defensin (Cremycin-5) for increased potency and stability.
Main Methods:
- Sequence-activity comparison between Cremycin-5 and paralogs to identify key sites.
- Saturation mutagenesis at identified sites to screen for enhanced activity.
- Antifungal activity assays against multiple fungal species, including Candida albicans clinical isolates.
- Molecular dynamic simulations to elucidate the mechanism of action.
- Analysis of sequence, structure, and epistasis for evolutionary insights.
Main Results:
- Identified a Universally Enhanceable Activity-Modulating Site (UEAMS) at position Glu-15.
- Mutations at Glu-15 universally enhanced antifungal activity against multiple fungal species.
- The Glu15Lys mutant showed >9-fold increased potency against Candida albicans, inhibiting cytokinesis.
- The engineered mutant exhibited high stability, rapid killing kinetics, and no detectable hemolysis.
- Molecular dynamics suggested allosteric regulation of distant functional residues via UEAMS mutations.
Conclusions:
- The evolution-based strategy successfully engineered a potent antifungal AMP.
- UEAMS represents a novel target for improving AMP efficacy and overcoming limitations.
- The findings provide a new pathway for developing AMP-based antifungal drugs.
- Understanding epistatic interactions can guide future AMP engineering efforts.
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