Methods of isolation and purification of outer membrane vesicles from gram-negative bacteria

Klimentová, Jana; Stulı́k, Jiřı́

doi:10.1016/j.micres.2014.09.006

Cited by 237 publications

(234 citation statements)

References 78 publications

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“…Consequently, both membranes have to remain functional even under variable conditions. The bestinvestigated mechanism that involves changes in the outer membrane of Gram-negative bacteria is the formation of outer membrane vesicles (OMVs) (Kulp & Kuehn, 2010;Klimentová & Stulík, 2015). The OMVs are released during all growth phases of the bacterial cultures (Kadurugamuwa & Beveridge, 1995;Manning & Kuehn, 2011), although their amount and composition may vary according to the growing conditions.…”

Section: Introductionmentioning

confidence: 99%

“…Structural studies of OMV invariably showed that they are small spherical particles, from 50 to 250 nm, enclosed by a membrane bilayer, that contain phospholipids (PLs), LPSs, proteins and in some cases nucleic acids. They are also involved in the transport of metabolites, toxins, signalling molecules and other small molecules (Horstman & Kuehn, 2000Dutta et al, 2004;Keenan et al, 2008;Bomberger et al, 2009;Schertzer & Whiteley, 2012;Biller et al, 2014;Klimentová & Stulík, 2015). Furthermore, the formation of OMVs is a crucial part of the bacterial envelope stress response, protecting the bacteria from internal and external stressors (Manning & Kuehn, 2011;Baumgarten et al, 2012;MacDonald & Kuehn, 2012;Schwechheimer et al, 2013Schwechheimer et al, , 2014.…”

Section: Introductionmentioning

confidence: 99%

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Release of outer membrane vesicles in Pseudomonas putida as a response to stress caused by cationic surfactants

et al. 2016

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Release of outer membrane vesicles inPseudomonas putida A (ATCC 12633), a degrader of cationic surfactants, releases outer membrane vesicles (OMVs) when grown with tetradecyltrimethylammonium bromide (TTAB) as the sole carbon, nitrogen and energy source. The OMVs exhibit a bilayer structure and were found to be composed of lipopolysaccharides, proteins and phospholipids (PLs) such as cardiolipin, phosphatidylcholine, phosphatidic acid and phosphatidylglycerol (PG). The OMVs showed a marked increase in the PG content, approximately 43 % higher than the amount registered in the parent cells from which the vesicles were derived. After growth of P. putida with TTAB, the amount of lipoprotein covalently cross-linked to the peptidoglycan showed a twofold decrease when compared with values found after growth without the surfactant [16¡2 and 28¡3 mg (mg cell envelope protein) 21, respectively]. This decrease in the amount of lipoprotein can be related to areas of loss of contact between the outer membrane and the peptidoglycan and, therefore, to OMV production. In addition, due to its amphiphilic nature, TTAB can contribute to OMV biogenesis, through a physical mechanism, by induction of the curvature of the membrane. Taking into account that OVMs were produced when the cells were grown under external stress, caused by the surfactant, and that TTAB was detected in the vesicles [48 nmol TTAB (nmol PL) 21], we concluded that this system of TTAB elimination is a mechanism that P. putida A (ATCC 12633) would utilize for alleviating stress caused by cationic surfactants.

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Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Release of outer membrane vesicles in Pseudomonas putida as a response to stress caused by cationic surfactants

et al. 2016

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“…Methods used for the isolation of the OMV and EV include ultracentrifugation and precipitation [2]. The isolation of OMV starts from culture supernatant after removal of cultured bacteria by centrifugation at 6,000 × g for 5 min.…”

Section: Isolation Purification and Quantification Of Omv And Evmentioning

confidence: 99%

“…OMVs are also reported to behave as secretion system for factors related to virulence and toxins inside a host, facilitate survival in the inter-species environmental competition, intra-and inter-species communication, secretion of proteins which are misfolded and aggregated (a novel stress response mechanism) etc. [2]. Both pathogenic and non-pathogenic species of Gram-negative bacteria are known to release vesicles [3][4][5][6], including Escherichia coli [7,8], Borrelia burgdorferi [9], Neisseria spp.…”

Section: Introductionmentioning

confidence: 99%

Significances of OMV and Extracellular Vesicle Proteomics

Tiwari¹

2017

J Data Mining Genomics Proteomics

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Outer Membrane Vesicle (OMV) proteome has been involved into the pathogenesis of diseases and resistance of microorganisms against a number of antibiotics, mechanism of action of probiotics and host-pathogen interaction etc. We have enlightened the role played by extracellular vesicles proteomics in the pathogenesis of different diseases related to human. Isolation succeeded by purification of ample amount of OMV from biological samples, is one of the most important steps for further proteome related analysis. With the development of both labelled and label-free methods used in proteomics, significant progress has been made in previous years in membrane proteomics. Hence, it is important to review the biological significance of proteins found in the OMV fractions using membrane proteomics approach. We have also explained methods used for isolation, purification and quantification of OMV. In the present review, it can be concluded that proteomic study of outer membrane and extracellular vesicles has now gained priority for the detailed study of disease pathogenesis, drug resistance, vaccine development, cell signalling etc.

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“…Most commonly, EVs/OMVs are purified via ultracentrifugation, density gradient fractionation, immunoaffinity, precipitation or ultrafiltration techniques [15]. Ultracentrifugation, which is utilized for both pelleting EV/OMV and density gradient fractionation, requires centrifugal forces of 100,000 × g or greater, necessitating access to expensive and specialized centrifugation equipment.…”

Section: Introductionmentioning

confidence: 99%

Affinity purification of bacterial outer membrane vesicles (OMVs) utilizing a His-tag mutant

Alves

Turner

DiVito

et al. 2017

Research in Microbiology

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To facilitate the rapid purification of bacterial outer membrane vesicles (OMVs), we developed two plasmid constructs that utilize a truncated, transmembrane protein to present an exterior histidine repeat sequence. We chose OmpA, a highly abundant porin protein, as the protein scaffold and utilized the lac promoter to allow for inducible control of the epitope-presenting construct. OMVs containing mutant OmpA-His6 were purified directly from Escherichia coli culture media on an immobilized metal affinity chromatography (IMAC) Ni-NTA resin. This enabling technology can be combined with other molecular tools directed at OMV packaging to facilitate the separation of modified/cargo-loaded OMV from their wt counterparts. In addition to numerous applications in the pharmaceutical and environmental remediation industries, this technology can be utilized to enhance basic research capabilities in the area of elucidating endogenous OMV function.

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Methods of isolation and purification of outer membrane vesicles from gram-negative bacteria

Cited by 237 publications

References 78 publications

Release of outer membrane vesicles in Pseudomonas putida as a response to stress caused by cationic surfactants

Release of outer membrane vesicles in Pseudomonas putida as a response to stress caused by cationic surfactants

Significances of OMV and Extracellular Vesicle Proteomics

Affinity purification of bacterial outer membrane vesicles (OMVs) utilizing a His-tag mutant

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