In Vivo Bactericidal Efficacy of GWH1 Antimicrobial Peptide Displayed on Protein Nanoparticles, a Potential Alternative to Antibiotics.

François Malouin,Neus Ferrer-Miralles,Antonio Villaverde,Eric Brouillette,Jose V Carratalá,Naroa Serna,Alejandro Sánchez-Chardi,Elena Garcia-Fruitós,Anna Arís,Julieta M Sánchez

doi:10.3390/pharmaceutics12121217

Abstract

Oligomerization of antimicrobial peptides into nanosized supramolecular complexes produced in biological systems (inclusion bodies and self-assembling nanoparticles) seems an appealing alternative to conventional antibiotics. In this work, the antimicrobial peptide, GWH1, was N-terminally fused to two different scaffold proteins, namely, GFP and IFN-γ for its bacterial production in the form of such recombinant protein complexes. Protein self-assembling as regular soluble protein nanoparticles was achieved in the case of GWH1-GFP, while oligomerization into bacterial inclusion bodies was reached in both constructions. Among all these types of therapeutic proteins, protein nanoparticles of GWH1-GFP showed the highest bactericidal effect in an in vitro assay against Escherichia coli, whereas non-oligomerized GWH1-GFP and GWH1-IFN-γ only displayed a moderate bactericidal activity. These results indicate that the biological activity of GWH1 is specifically enhanced in the form of regular multi-display configurations. Those in vitro observations were fully validated against a bacterial infection using a mouse mastitis model, in which the GWH1-GFP soluble nanoparticles were able to effectively reduce bacterial loads.

Highlights

Novel antimicrobial and engineering approaches are urgently needed to cope with antibiotic resistance
The extensive use of antibiotics for the treatment and prevention of bacterial infections increases the selective pressure for appearance of resistant bacteria
We have described the antimicrobial activity of peptides fused to a scaffold protein and displayed in various copies on soluble protein-only nanoparticles (PNPs)

Summary

Introduction

Novel antimicrobial and engineering approaches are urgently needed to cope with antibiotic resistance. In 2017, Serna et al described for the first time the use of PNPs as antibacterial agents [7] These soluble PNPs were obtained following a modular protein design based on the fusion of a cationic peptide to a C-terminal his-tagged scaffold protein [8]. Bacterial IBs, once considered as waste by-products derived from recombinant protein production, provide a useful source of ready-to-use active protein Inside these structures, therapeutic proteins are stored in native and native-like conformations and are released under physiological conditions [12,13,14]. Therapeutic proteins are stored in native and native-like conformations and are released under physiological conditions [12,13,14] The benefits of this system lie in the protective effect against degradation and the sustained release of the protein, which in both cases can significantly increase their half-lives

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