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Tick saliva contains a complex mix of anti\u2010host defense factors, including the immunosuppressive cysteine\u2010rich secretory glycoprotein Salp15 from<jats:italic>Ixodes scapularis<\/jats:italic> ticks and orthologs like Iric\u20101 from<jats:italic>Ixodes<\/jats:italic><jats:italic>ricinus<\/jats:italic>. All tick\u2010borne microbes benefit from the immunosuppression at the tick bite site; in addition, borreliae exploit the binding of Salp15 to their outer surface protein C (OspC) for enhanced transmission. Hence, Salp15 proteins are attractive targets for anti\u2010tick vaccines that also target borreliae. However, recombinant Salp proteins are not accessible in sufficient quantity for either vaccine manufacturing or for structural characterization. As an alternative to low\u2010yield eukaryotic systems, we investigated cytoplasmic expression in<jats:italic>Escherichia coli<\/jats:italic>, even though this would not result in glycosylation. His\u2010tagged Salp15 was efficiently expressed but insoluble. Among the various solubility\u2010enhancing protein tags tested, DsbA was superior, yielding milligram amounts of soluble, monomeric Salp15 and Iric\u20101 fusions. Easily accessible mutants enabled epitope mapping of two monoclonal antibodies that, importantly, cross\u2010react with glycosylated Salp15, and revealed interaction sites with OspC. Free Salp15 and Iric\u20101 from protease\u2010cleavable fusions, despite limited solubility, allowed the recording of<jats:sup>1<\/jats:sup>H\u2013<jats:sup>15<\/jats:sup>N 2D NMR spectra, suggesting partial folding of the wild\u2010type proteins but not of Cys\u2010free variants. Fusion to the NMR\u2010compatible GB1 domain sufficiently enhanced solubility to reveal first secondary structure elements in<jats:sup>13<\/jats:sup>C\/<jats:sup>15<\/jats:sup>N double\u2010labeled Iric\u20101. Together,<jats:italic>E. coli<\/jats:italic> expression of appropriately fused Salp15 proteins may be highly valuable for the molecular characterization of the function and eventually the 3D structure of these medically relevant tick proteins.<\/jats:p>","DOI":"10.1016\/j.fob.2014.12.002","type":"journal-article","created":{"date-parts":[[2014,12,24]],"date-time":"2014-12-24T17:03:01Z","timestamp":1419440581000},"page":"42-55","update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":13,"title":["Soluble cysteine\u2010rich tick saliva proteins Salp15 and Iric\u20101 from<i>E. coli<\/i>"],"prefix":"10.1002","volume":"5","author":[{"given":"Philipp","family":"Kolb","sequence":"first","affiliation":[{"name":"University Hospital Freiburg, Internal Medicine 2\/Molecular Biology, Hugstetter Str. 55, D-79106 Freiburg, Germany"},{"name":"University of Freiburg, Biological Faculty, Sch\u00e4nzlestr. 1, D-79104 Freiburg, 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