Engineering a customizable antibacterial T6SS-based platform in<i>Vibrio natriegens</i>

Jana, Biswanath; Keppel, Kinga; Salomon, Dor

doi:10.1101/2021.05.04.439770

Cited by 2 publications

(1 citation statement)

References 52 publications

(79 reference statements)

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“…For example, T3SSs from diverse pathogenic bacteria were used as translocation tools to deliver proteins into mammalian cells ( 4 , 5 ). T6SS, another widespread injectisome in pathogens, was developed as delivery vector in biotechnological applications ( 30 , 31 ). Many other protein complexes, such as Tc toxins ( 10 , 11 ), type IV pilins ( 32 , 33 ) and pore-forming proteins ( 34 , 35 ), were also used to accomplish similar assignments toward eukaryotic cells, particularly killing cancer cells ( 36 ).…”

Section: Discussionmentioning

confidence: 99%

N-terminal signal peptides facilitate the engineering of PVC complex as a potent protein delivery system

et al. 2022

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Extracellular contractile injection systems (eCISs) are widespread bacterial nanomachines that resemble T4 phage tail. As a typical eCIS, Photorhabdus virulence cassette (PVC) was proposed to inject toxins into eukaryotic cells by puncturing the cell membrane from outside. This makes it an ideal tool for protein delivery in biomedical research. However, how to manipulate this nanocomplex as a molecular syringe is still undetermined. Here, we identify that one group of N-terminal signal peptide (SP) sequences are crucial for the effector loading into the inner tube of PVC complex. By application of genetic operation, cryo–electron microscopy, in vitro translocation assays, and animal experiments, we show that, under the guidance of the SP, numerous prokaryotic and eukaryotic proteins can be loaded into PVC to exert their functions across cell membranes. We therefore might customize PVC as a potent protein delivery nanosyringe for biotherapy by selecting cargo proteins in a broad spectrum, regardless of their species, sizes, and charges.

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Section: Discussionmentioning

confidence: 99%

N-terminal signal peptides facilitate the engineering of PVC complex as a potent protein delivery system

et al. 2022

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A genomic and structural bioinformatic pipeline identifies candidate type VI secretion antibacterial effector-immunity pairs

Geller

Zlotkin

Shalom

et al. 2023

Preprint

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Type VI secretion systems (T6SS) are common bacterial contractile injection systems that inject toxic "effector" proteins into neighboring cells. We bioinformatically investigated T6SS core proteins in 11,832 genomes of Gram negative bacteria. Comparison of T6SS core proteins that are covalently attached to toxic T6SS effector proteins (T6Es) versus those that are not revealed differences in phylogenetic distribution, physical properties, and genomic position. Using the data generated from our bioinformatic analysis, we developed a new genomic- and Alphafold2-based pipeline for discovery of putative T6Es. We experimentally validated the toxic and immunity activities of four putative antibacterial T6SS effector proteins and four cognate immunity genes from diverse species, respectively. We used Foldseek to predict possible mechanisms of action of the putative T6Es, which was much more effective than sequence-based methods. Evidence of the possible mechanisms of action of the putative T6Es was explored through fluorescence microscopy, where we observed cell wall-targeting, DNA degradation, and cell filamentation. This study shows how combining genomic data mining with new structure-based bioinformatic tools can facilitate identification of novel antibacterial toxins.

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Engineering a customizable antibacterial T6SS-based platform inVibrio natriegens

Cited by 2 publications

References 52 publications

N-terminal signal peptides facilitate the engineering of PVC complex as a potent protein delivery system

N-terminal signal peptides facilitate the engineering of PVC complex as a potent protein delivery system

A genomic and structural bioinformatic pipeline identifies candidate type VI secretion antibacterial effector-immunity pairs

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