{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,10,12]],"date-time":"2025-10-12T03:10:03Z","timestamp":1760238603783,"version":"build-2065373602"},"reference-count":33,"publisher":"MDPI AG","issue":"9","license":[{"start":{"date-parts":[[2020,8,20]],"date-time":"2020-08-20T00:00:00Z","timestamp":1597881600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Maryland Industrial Partnershipd","award":["6409"],"award-info":[{"award-number":["6409"]}]},{"DOI":"10.13039\/100000002","name":"National Institutes of Health","doi-asserted-by":"publisher","award":["GM21248","R43AI138612"],"award-info":[{"award-number":["GM21248","R43AI138612"]}],"id":[{"id":"10.13039\/100000002","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Pharmaceutics"],"abstract":"<jats:p>Identification of antigens is important for vaccine production. We tested extraction protocols using cetyltrimethylammonium tosylate (CTAT) and sodium dodecylbenzenesulfonate (SDBS) to formulate surfactant vesicles (SVs) containing components from Neisseria gonorrhoeae. Carbohydrate and protein assays demonstrated that protein and carbohydrates were incorporated into the vesicle leaflet. Depending on the extraction protocol utilized, 100\u2013400 \u00b5g of protein\/mL of SVs solution was obtained. Gel electrophoresis followed by silver staining demonstrated that SV extracts contained lipooligosaccharide and a subset of bacterial proteins and lipoproteins. Western blotting and mass spectral analysis indicated that the majority of the proteins were derived from the outer membrane. Mass spectrometric and bioinformatics analysis of SVs identified 29 membrane proteins, including porin and opacity-associated protein. Proteins embedded in the SVs leaflet could be degraded by the addition of trypsin or proteinase K. Our data showed that the incorporation of CTAT and SDBS into vesicles eliminated their toxicity as measured by a THP-1 killing assay. Incorporation of gonococcal cell surface components into SVs reduced toxicity as compared to the whole cell extracts, as measured by cytokine induction, while retaining the immunogenicity. This process constitutes a general method for extracting bacterial surface components and identification of antigens that might be included in vaccines.<\/jats:p>","DOI":"10.3390\/pharmaceutics12090787","type":"journal-article","created":{"date-parts":[[2020,8,20]],"date-time":"2020-08-20T21:13:42Z","timestamp":1597958022000},"page":"787","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":2,"title":["Extraction of Membrane Components from Neisseria gonorrhoeae Using Catanionic Surfactant Vesicles: A New Approach for the Study of Bacterial Surface Molecules"],"prefix":"10.3390","volume":"12","author":[{"given":"Daniel C.","family":"Stein","sequence":"first","affiliation":[{"name":"Department of Cell Biology and Molecular Genetics, University of Maryland, College Park, MD 20742, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Lenea","family":"H. Stocker","sequence":"additional","affiliation":[{"name":"Department of Chemistry and Biochemistry, University of Maryland, College Park, MD 20742, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Abigail E.","family":"Powell","sequence":"additional","affiliation":[{"name":"Department of Chemistry and Biochemistry, University of Maryland, College Park, MD 20742, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Salsawi","family":"Kebede","sequence":"additional","affiliation":[{"name":"Department of Cell Biology and Molecular Genetics, University of Maryland, College Park, MD 20742, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"David","family":"Watts","sequence":"additional","affiliation":[{"name":"SD Nanosciences, College Park, MD 20742, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Emma","family":"Williams","sequence":"additional","affiliation":[{"name":"Department of Cell Biology and Molecular Genetics, University of Maryland, College Park, MD 20742, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Nicholas","family":"Soto","sequence":"additional","affiliation":[{"name":"Department of Chemistry and Biochemistry, University of Maryland, College Park, MD 20742, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0799-3223","authenticated-orcid":false,"given":"Avantika","family":"Dhabaria","sequence":"additional","affiliation":[{"name":"Department of Chemistry and Biochemistry, University of Maryland, College Park, MD 20742, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4979-0229","authenticated-orcid":false,"given":"Catherine","family":"Fenselau","sequence":"additional","affiliation":[{"name":"Department of Chemistry and Biochemistry, University of Maryland, College Park, MD 20742, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1955-2772","authenticated-orcid":false,"given":"Shweta","family":"Ganapati","sequence":"additional","affiliation":[{"name":"SD Nanosciences, College Park, MD 20742, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Philip","family":"DeShong","sequence":"additional","affiliation":[{"name":"Department of Chemistry and Biochemistry, University of Maryland, College Park, MD 20742, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2020,8,20]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"1220","DOI":"10.1126\/science.1175668","article-title":"Cell signaling in space and time: Where proteins come together and when they\u2019re apart","volume":"326","author":"Scott","year":"2009","journal-title":"Science"},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"633","DOI":"10.1016\/S0955-0674(03)00099-1","article-title":"Bacterial pathogenesis: Exploiting cellular adherence","volume":"15","author":"Boyle","year":"2003","journal-title":"Curr. Opin. Cell Biol."},{"key":"ref_3","first-page":"95","article-title":"Architecture and physiology of microbial biofilms","volume":"69","author":"Lazar","year":"2010","journal-title":"Roum. Arch. Microbiol. Immunol."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"753","DOI":"10.1590\/S0100-879X2002000700001","article-title":"Membrane proteins: Functional and structural studies using reconstituted proteoliposomes and 2-D crystals","volume":"35","author":"Rigaud","year":"2002","journal-title":"Braz. J. Med. Biol. Res."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"102","DOI":"10.1186\/1556-276X-8-102","article-title":"Liposome: Classification, preparation, and applications","volume":"8","author":"Akbarzadeh","year":"2013","journal-title":"Nanoscale Res. Lett."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"1916","DOI":"10.1039\/b806059b","article-title":"Carbohydrate-Functionalized Catanionic Surfactant Vesicles: Preparation and Lectin Binding Studies","volume":"4","author":"Park","year":"2008","journal-title":"Soft Matter"},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"5471","DOI":"10.1021\/ja8076439","article-title":"Carbohydrate Modified Catanionic Vesicles: Probing Multivalent Binding at the Bilayer Interface","volume":"131","author":"Thomas","year":"2009","journal-title":"J. Am. Chem. Soc."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"949","DOI":"10.1084\/jem.191.6.949","article-title":"Role of lipooligosaccharide in opa-independent invasion of Neisseria gonorrhoeae into human epithelial cells","volume":"191","author":"Song","year":"2000","journal-title":"J. Exp. Med."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"762","DOI":"10.1128\/iai.56.4.762-765.1988","article-title":"Use of transformation to construct Neisseria gonorrhoeae strains with altered lipooligosaccharides","volume":"56","author":"Stein","year":"1988","journal-title":"Infect. Immun."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"76","DOI":"10.1016\/0003-2697(85)90442-7","article-title":"Measurement of protein using bicinchoninic acid","volume":"150","author":"Smith","year":"1985","journal-title":"Anal. Biochem."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"1852","DOI":"10.1038\/nprot.2006.288","article-title":"Silver staining of proteins in polyacrylamide gels","volume":"1","author":"Chevallet","year":"2006","journal-title":"Nat. Protoc."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"4350","DOI":"10.1073\/pnas.76.9.4350","article-title":"Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: Procedure and some applications","volume":"76","author":"Towbin","year":"1979","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"6468","DOI":"10.1128\/JB.00969-12","article-title":"Construction and characterization of a derivative of Neisseria gonorrhoeae strain MS11 devoid of all opa genes","volume":"194","author":"LeVan","year":"2012","journal-title":"J. Bacteriol."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"2930","DOI":"10.1128\/jb.176.10.2930-2937.1994","article-title":"Role of phosphoglucomutase in lipooligosaccharide biosynthesis in Neisseria gonorrhoeae","volume":"176","author":"Sandlin","year":"1994","journal-title":"J. Bacteriol."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"2856","DOI":"10.1038\/nprot.2006.468","article-title":"In-gel digestion for mass spectrometric characterization of proteins and proteomes","volume":"1","author":"Shevchenko","year":"2006","journal-title":"Nat. Protoc."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"1134","DOI":"10.1021\/pr301107x","article-title":"Enrichment of plasma membrane proteins using nanoparticle pellicles: Comparison between silica and higher density nanoparticles","volume":"12","author":"Choksawangkarn","year":"2013","journal-title":"J. Proteome Res."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"801","DOI":"10.1146\/annurev.anchem.1.031207.112747","article-title":"Liposomes: Technologies and Analytical Applications","volume":"1","author":"Jesorka","year":"2008","journal-title":"Ann. Rev. Anal. Chem."},{"key":"ref_18","first-page":"49","article-title":"Clinical development of liposome-based drugs: Formulation, characterization, and therapeutic efficacy","volume":"7","author":"Chang","year":"2012","journal-title":"Int. J. Nanomed."},{"key":"ref_19","first-page":"1","article-title":"Liposomes as vaccine delivery systems: A review of the recent advances","volume":"2","author":"Schwendener","year":"2014","journal-title":"Ther. Adv. Vaccines"},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"476","DOI":"10.1016\/0005-2736(88)90094-6","article-title":"Purification of membrane proteins in SDS and subsequent renaturation","volume":"939","author":"Hjerten","year":"1988","journal-title":"Biochim. Biophys. Acta"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"302","DOI":"10.1016\/0021-9797(90)90348-R","article-title":"On the thermodynamic stability of liposomes","volume":"140","author":"Lasic","year":"1990","journal-title":"J. Colloid Interface Sci."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"82","DOI":"10.1016\/0005-2736(69)90061-3","article-title":"Potassium permeability of single compartment liposomes with and without valinomycin","volume":"193","author":"Johnson","year":"1969","journal-title":"Biochim. Biophys. Acta"},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"975","DOI":"10.2147\/IJN.S68861","article-title":"Liposomes as nanomedical devices","volume":"10","author":"Bozzuto","year":"2015","journal-title":"Int. J. Nanomed."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"261","DOI":"10.1146\/annurev-physchem-040215-112634","article-title":"Phospholipid Bilayers: Stability and Encapsulation of Nanoparticles","volume":"68","author":"Alipour","year":"2017","journal-title":"Annu. Rev. Phys. Chem."},{"key":"ref_25","unstructured":"Philippot, J., and Schuber, F. (1994). Liposomes as Tools for the Reconstitution of Biological Systems. Liposomes as Tools in Basic Research and Industry, CRC Press."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"419","DOI":"10.1529\/biophysj.104.040360","article-title":"A new method for the reconstitution of membrane proteins into giant unilamellar vesicles","volume":"87","author":"Girard","year":"2004","journal-title":"Biophys. J."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"1589","DOI":"10.3390\/ijms14011589","article-title":"Reconstitution of membrane proteins into model membranes: Seeking better ways to retain protein activities","volume":"14","author":"Shen","year":"2013","journal-title":"Int. J. Mol. Sci."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"1371","DOI":"10.1126\/science.2781283","article-title":"Spontaneous vesicle formation in aqueous mixtures of single-tailed surfactants","volume":"245","author":"Kaler","year":"1989","journal-title":"Science (New York, NY )"},{"key":"ref_29","unstructured":"Stocker, L. (2013). Catanionic SUrfactant Vesicles: Technology for Vaccine Depvelopment and Targetted Drug Delivery Applications. [Ph.D. Thesis, University of Maryland]."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"438","DOI":"10.21037\/atm.2016.08.53","article-title":"THP-1 cells as a model for human monocytes","volume":"4","author":"Bosshart","year":"2016","journal-title":"Ann. Transl. Med."},{"key":"ref_31","first-page":"491","article-title":"Macrophage cytokines: Involvement in immunity and infectious diseases","volume":"5","author":"Descoteaux","year":"2014","journal-title":"Front. Immunol."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"5771","DOI":"10.4049\/jimmunol.180.9.5771","article-title":"IL-10: The master regulator of immunity to infection","volume":"180","author":"Couper","year":"2008","journal-title":"J. Immunol."},{"key":"ref_33","doi-asserted-by":"crossref","unstructured":"Vijayan, V., Mohapatra, A., Uthaman, S., and Park, I.K. (2019). Recent Advances in Nanovaccines Using Biomimetic Immunomodulatory Materials. Pharmaceutics, 11.","DOI":"10.3390\/pharmaceutics11100534"}],"container-title":["Pharmaceutics"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1999-4923\/12\/9\/787\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T10:03:40Z","timestamp":1760177020000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1999-4923\/12\/9\/787"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2020,8,20]]},"references-count":33,"journal-issue":{"issue":"9","published-online":{"date-parts":[[2020,9]]}},"alternative-id":["pharmaceutics12090787"],"URL":"https:\/\/doi.org\/10.3390\/pharmaceutics12090787","relation":{},"ISSN":["1999-4923"],"issn-type":[{"type":"electronic","value":"1999-4923"}],"subject":[],"published":{"date-parts":[[2020,8,20]]}}}