Affinity purification of bacterial outer membrane vesicles (OMVs) utilizing a His-tag mutant |
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| Affiliation: | 1. National Research Council, 500 Fifth Street NW, Keck 576, Washington, DC 20001, USA;2. Department of Emergency Medicine, Indiana University School of Medicine, Indianapolis, IN 46202, USA;3. Center for Bio/Molecular Science & Engineering, Naval Research Laboratory, Washington, DC 20375, USA;4. American Society for Engineering Education (ASEE), 1818 N Street NW, Suite 600, Washington, DC 20036, USA;5. North Carolina State University, Joint Department of Biomedical Engineering, UNC-Chapel Hill/NC State University, 2068 Engineering Building 2, Campus Box 7911, Raleigh, NC 27695, USA;1. Department of polymers Material Research, Institute of Advanced Technology and New Materials, City for Scientific Research and Technology Applications, New Borg El-Arab City 21934, Alexandria, Egypt;2. Department of Chemistry, Faculty of Science, Zagazig University, Zagazig, Egypt;1. Laboratory of Pediatric Infectious Diseases, Department of Pediatrics, Radboud Institute for Molecular Life Sciences, Radboud University Medical Center, Nijmegen, The Netherlands;2. Section Molecular Microbiology, Department of Molecular Cell Biology, Faculty of Earth and Life Sciences, VU University, Amsterdam, The Netherlands;3. Abera Bioscience AB, Stockholm, Sweden;1. Department of Life Science, Division of Molecular and Life Sciences, Pohang University of Science and Technology, Pohang, Gyeongsangbuk-do 37673, South Korea;2. Department of Chemistry, Chemistry Institute for Functional Materials, Pusan National University, Busan 46241, South Korea;1. Meinig School of Biomedical Engineering, Cornell University, Ithaca, NY 14853, USA;2. Department of Biological and Environmental Engineering, Cornell University, Ithaca, NY 14853, USA;3. School of Chemical and Biomolecular Engineering, Cornell University, Ithaca, NY 14853, USA;4. Department of Microbiology and Immunology, Cornell University, Ithaca, NY 14853, USA;1. Institute of Molecular Biosciences, University of Graz, NAWI Graz, BioTechMed-Graz, Humboldtstraße 50, A-8010 Graz, Austria;2. Institute of Biochemistry, Graz University of Technology, NAWI Graz, BioTechMed-Graz, Petersgasse 12/2, A-8010 Graz, Austria;3. Institute of Chemistry/Organic and Bioorganic Chemistry, University of Graz, NAWI Graz, BioTechMed-Graz, Heinrichstraße 28, A-8010 Graz, Austria;4. Alberta Glycomics Centre, Department of Biological Sciences, University of Alberta, CW405 Biological Sciences Building, Edmonton, AB, Canada T6G 2E9;5. Institute of Biophysics, Medical University of Graz, BioTechMed-Graz, Harrachgasse 21, A-8010 Graz, Austria |
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| Abstract: | ![]()
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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| Keywords: | Outer membrane vesicles (OMVs) Extracellular vesicles Affinity purification IMAC His-tag |
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