A modular platform for on-demand vaccine self-assembly enabled by decoration of bacterial outer membrane vesicles with biotinylated antigens

AbstractEngineered outer membrane vesicles (OMVs) derived from laboratory strains of bacteria are a promising technology for the creation of non-infectious, nanoparticle vaccines against diverse pathogens. As mimics of the bacterial cell surface, OMVs offer a molecularly-defined architecture for programming repetitive, high-density display of heterologous antigens in conformations that elicit strong B and T cell immune responses. However, antigen display on the surface of OMVs can be difficult to control and highly variable due to bottlenecks in protein expression and localization to the outer membrane of the host cell, especially for bulky and/or complex antigens. To address this shortcoming, we created a universal approach called AddVax (avidin-based dock- and-display for vaccine antigen cross (x)-linking) whereby virtually any antigen that is amenable to biotinylation can be linked to the exterior of OMVs whose surfaces are remodeled with multiple copies of a synthetic antigen receptor (SNARE) comprised of an outer membrane scaffold protein fused to a member of the avidin family. We show that SNARE-OMVs can be readily decorated with a molecularly diverse array of biotinylated subunit antigens, including globular and membrane proteins, glycans and glycoconjugates, haptens, lipids, and short peptides. When the resulting OMV formulations were injected in wild-type BALB/c mice, strong antigen-specific antibody responses were observed that depended on the physical coupling between the antigen and SNARE-OMV delivery vehicle. Overall, these results demonstrate AddVax as a modular platform for rapid self-assembly of antigen-studded OMVs with the potential to accelerate vaccine generation, respond rapidly to pathogen threats in humans and animals, and simplify vaccine stockpiling.

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Surface Modification of E. coli Outer Membrane Vesicles with Glycosylphosphatidylinositol-Anchored Proteins: Generating Pro/Eukaryote Chimera Constructs

Membranes ◽

10.3390/membranes11060428 ◽

2021 ◽

Vol 11 (6) ◽

pp. 428

Author(s):

Marianne Zaruba ◽

Lena Roschitz ◽

Haider Sami ◽

Manfred Ogris ◽

Wilhelm Gerner ◽

...

Keyword(s):

Surface Modification ◽

Outer Membrane ◽

Extracellular Vesicles ◽

Membrane Vesicles ◽

Outer Membrane Vesicles ◽

Functional Surface ◽

Vaccine Strategy ◽

Fluorescent Marker ◽

E Coli ◽

Modular Platform

Extracellular vesicles produced by different types of cells have recently attracted great attention, not only for their role in physiology and pathology, but also because of the emerging applications in gene therapy, vaccine production and diagnostics. Less well known than their eukaryotic counterpart, also bacteria produce extracellular vesicles, in the case of the Gram-negative E. coli the main species is termed outer membrane vesicles (OMVs). In this study, we show for the first time the functional surface modification of E. coli OMVs with glycosylphosphatidylinositol (GPI)-anchored protein, exploiting a process variably described as molecular painting or protein engineering in eukaryotic membranes, whereby the lipid part of the GPI anchor inserts in cell membranes. By transferring the process to bacterial vesicles, we can generate a hybrid of perfectly eukaryotic proteins (in terms of folding and post-translational modifications) on a prokaryotic platform. We could demonstrate that two different GPI proteins can be displayed on the same OMV. In addition to fluorescent marker proteins, cytokines, growth factors and antigens canb be potentially transferred, generating a versatile modular platform for a novel vaccine strategy.

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LOW-DOSE RECOMBINANT VACCINE ANTIGEN DELIVERY BY ENGINEERED OUTER MEMBRANE VESICLES

Nano LIFE ◽

10.1142/s1793984413420026 ◽

2013 ◽

Vol 03 (03) ◽

pp. 1342002

Author(s):

JOSEPH ROSENTHAL ◽

TIFFANY LEUNG ◽

MATTHEW DELISA ◽

DAVID PUTNAM

Keyword(s):

Injection Site ◽

Outer Membrane ◽

Immune Cell ◽

Membrane Vesicle ◽

Membrane Vesicles ◽

Outer Membrane Vesicles ◽

Vaccine Antigen ◽

Response Analysis ◽

Antigen Delivery ◽

Applied Dose

The use of engineered antigen carriers to optimize the immune response to recombinant subunit vaccines has seen great advances in recent years. Optimization can take several forms, such as facilitating stimulation of certain immune cells or amplifying the adjuvancy effect of the vaccine formulation. In this paper, we applied dose/response analysis to demonstrate the ability of outer membrane vesicle (OMV) antigen carriers derived from engineered Escherichia coli to produce strong antigen-specific immune responses to a model antigen at a significantly decreased antigen load compared to an industry standard alum-based control. Inflammopathology and histological analysis of extended studies further supported a capacity to enhance immune cell recruitment locally at the injection site while decreasing inflammation and eliminating injection site scaring. The results indicate a strong potential for OMV-based vaccines as recombinant antigen delivery vehicles, affording strong immunogenicity at low doses with a broadly applicable platform for recombinant subunit antigen inclusion.

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The Effect of Helicobacter Pylori Outer Membrane Vesicles On Vacuolation in Gastric and Colonic Epithelial Cells

Endoscopy ◽

10.1055/s-2006-956848 ◽

2006 ◽

Vol 38 (11) ◽

Author(s):

EM Mullaney ◽

AM Terres ◽

DP Kelleher

Keyword(s):

Helicobacter Pylori ◽

Epithelial Cells ◽

Outer Membrane ◽

Membrane Vesicles ◽

Outer Membrane Vesicles ◽

Colonic Epithelial Cells

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Electrophysiological Characterization of Transport Across Outer Membrane Channels from Gram-Negative Bacteria in Their Native Environment

10.26434/chemrxiv.8282204.v1 ◽

2019 ◽

Author(s):

Jiajun Wang ◽

Rémi Terrasse ◽

Jayesh Arun Bafna ◽

Lorraine Benier ◽

Mathias Winterhalter

Keyword(s):

Outer Membrane ◽

Lipid Membrane ◽

Membrane Vesicles ◽

Outer Membrane Vesicles ◽

Multi Drug Resistance ◽

Gram Negative Bacteria ◽

Membrane Channels ◽

Gram Negative ◽

Electrophysiological Characterization

Multi-drug resistance in Gram-negative bacteria is often associated with low permeability of the outer membrane. To investigate the role of membrane channels in the uptake of antibiotics, we extract, purify and reconstitute them into artificial planar membranes. To avoid this time-consuming procedure, here we show a robust approach using fusion of native outer membrane vesicles (OMV) into planar lipid bilayer which moreover allows also to some extend the characterization of membrane protein channels in their native environment. Two major membrane channels from Escherichia coli, OmpF and OmpC, were overexpressed from the host and the corresponding OMVs were collected. Each OMV fusion revealed surprisingly single or only few channel activities. The asymmetry of the OMV´s translates after fusion into the lipid membrane with the LPS dominantly present at the side of OMV addition. Compared to conventional reconstitution methods, the channels fused from OMVs containing LPS have similar conductance but a much broader distribution. The addition of Enrofloxacin on the LPS side yields somewhat higher association (kon) and lower dissociation (koff) rates compared to LPS-free reconstitution. We conclude that using outer membrane vesicles is a fast and easy approach for functional and structural studies of membrane channels in the native membrane.

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