Journal
RESEARCH IN MICROBIOLOGY
Volume 168, Issue 2, Pages 139-146Publisher
ELSEVIER SCIENCE BV
DOI: 10.1016/j.resmic.2016.10.001
Keywords
Escherichia coli; Outer membrane vesicles (OMVs); Extracellular vesicles; Affinity purification; IMAC; His-tag
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Funding
- Office of Naval Research through Core funds [MA041-06-41-T079-17]
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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/cargoloaded 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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