Exploiting phage receptor binding proteins to enable endolysins to kill Gram-negative bacteria.

Scientific Reports
Athina ZamparaLone Brøndsted

Abstract

Bacteriophage-encoded endolysins degrading the bacterial peptidoglycan are promising antibacterials for combating antibiotic-resistant bacteria. However, endolysins have limited use against Gram-negative bacteria, since the outer membrane prevents access to the peptidoglycan. Here, we present Innolysins, an innovative concept for engineering endolysins to exert antibacterial activity against Gram-negative bacteria. Innolysins combine the enzymatic activity of endolysins with the binding capacity of phage receptor binding proteins (RBPs). As proof-of-concept, we constructed 12 Innolysins by fusing phage T5 endolysin and RBP Pb5 in different configurations. One of these, Innolysin Ec6 displayed antibacterial activity against Escherichia coli only in the presence of Pb5 receptor FhuA, leading to 1.22 ± 0.12 log reduction in cell counts. Accordingly, other bacterial species carrying FhuA homologs such as Shigella sonnei and Pseudomonas aeruginosa were sensitive to Innolysin Ec6. To enhance the antibacterial activity, we further constructed 228 novel Innolysins by fusing 23 endolysins with Pb5. High-throughput screening allowed to select Innolysin Ec21 as the best antibacterial candidate, leading to 2.20 ± 0.09 log reduction in E. c...Continue Reading

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Citations

Jan 17, 2021·International Journal of Molecular Sciences·Anna Allué-GuardiaJordi B Torrelles
Oct 15, 2020·Current Opinion in Biotechnology·Diana Gutiérrez, Yves Briers
Feb 19, 2021·Frontiers in Microbiology·Athina ZamparaLone Brøndsted
Mar 14, 2021·Current Opinion in Biotechnology·Joana AzeredoZuzanna Drulis-Kawa
Jun 26, 2021·Infectious Diseases and Therapy·Bilal AslamZulqarnain Baloch
Jul 3, 2021·Antibiotics·Pablo Cristobal-CuetoMeritxell García-Quintanilla
Jul 23, 2021·Research in Microbiology·Bibi Fathima, Ann Catherine Archer

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Methods Mentioned

BETA
Transmission electron microscopy
Protein Assay

Software Mentioned

GraphPad Prism
iTEM
Clustal Omega
Jalview
BLASTP

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