Improved smallest peptides based on positive charge increase of the γ-core motif from PνD₁ and their mechanism of action against Candida species

Abstract

Background

Plant defensins have a hallmark γ-core motif (GXCX_3-9C) that is related to their antimicrobial properties. The aim of this work was to design synthetic peptides based on the region corresponding to the PvD₁ defensin γ-core that are the smallest amino acid sequences that bear the strongest biological activity.

Methods

We made rational substitutions of negatively charged amino acid residues with positively charged ones, and the reduction in length in the selected PvD₁ γ-core sequence to verify whether the increased net positive charges and shortened length are related to the increase in antifungal activity. Herein, we opted to evaluate the action mechanism of γ_33-41PvD₁⁺⁺ peptide due to its significant inhibitory effect on tested yeasts. In addition, it is the smallest construct comprising only nine amino acid residues, giving it a better possibility to be a prototype for designing a new antifungal drug, with lower costs to the pharmaceutical industry while still maintaining the strongest antimicrobial properties.

Results

The γ_33-41PvD₁⁺⁺ peptide caused the most toxic effects in the yeast Candida buinensis, leading to membrane permeabilization, viability loss, endogenous reactive oxygen species increase, the activation of metacaspase, and the loss of mitochondrial functionality, suggesting that this peptide triggers cell death via apoptosis.

Conclusion

We observed that the antifungal activity of PvD₁ is not strictly localized in the structural domain, which comprises the γ-core region and that the increase in the net positive charge is directly related to the increase in antifungal activity.

Keywords:

• defensin
• antimicrobial peptide
• cationic
• mechanism of action

Acknowledgments

This work was performed at the Universidade Estadual do Norte Fluminense Darcy Ribeiro. This study was financed in part by the Coordenação de Aperfeiçoamento de Pessoal de Nível Superior – Brasil(CAPES) – Finance Code 001. We acknowledge the financial support from the Brazilian agencies CNPq(305766/2013-9) FAPERJ (E-26/203090/2016; E-26/202.132/2015; E-26/202.735/2016). We are grateful to Souza LCD and Kokis VM for general laboratory technical support.

Author contributions

All authors contributed to data analysis, drafting and revising the article, gave final approval of the version to be published, and agree to be accountable for all aspects of the work.

Disclosure

The authors report no conflicts of interest in this work.