Recombinant protein production and purification of SiiD, SiiE and SiiF - Components of the SPI4-encoded type I secretion system from Salmonella Typhimurium

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dc.contributor.authorKlingl, Stefan
dc.contributor.authorKordes, Sina
dc.contributor.authorSchmid, Benedikt
dc.contributor.authorGerlach, Roman G.
dc.contributor.authorHensel, Michael
dc.contributor.authorMuller, Yves A.
dc.date.accessioned2021-12-23T16:16:29Z-
dc.date.available2021-12-23T16:16:29Z-
dc.date.issued2020
dc.identifier.issn10465928
dc.identifier.urihttps://osnascholar.ub.uni-osnabrueck.de/handle/unios/11898-
dc.description.abstractIn humans, Salmonella enterica infections are responsible for a plethora of medical conditions. These include intestinal inflammation and typhoid fever. The initial contact between Salmonella and polarized epithelial cells is established by the SPI4-encoded type I secretion system (T1SS), which secretes SiiE, a giant non-fimbrial adhesin. We have recombinantly produced various domains of this T1SS from Salmonella enterica serovar Typhimurium in Escherichia coli for further experimental characterization. We purified three variants of SiiD, the periplasmic adapter protein spanning the space between the inner and outer membrane, two variants of the SiiE N-terminal region and the N-terminal domain of the SiiF ATP-binding cassette (ABC) transporter. In all three proteins, at least one variant yielded high amounts of pure soluble protein. Secondary structure content and cooperative unfolding were investigated by circular dichroism (CD) spectroscopy. Secondary structure contents were in good agreement with estimates derived from SiiD and SiiF homology models or, in case of the SiiE N-terminal region, a secondary structure prediction. For one SiiD variant, protein crystals could be obtained that diffracted X-rays to approximately 4 angstrom resolution.
dc.description.sponsorshipDeutsche ForschungsgemeinschaftGerman Research Foundation (DFG) [HE1964/13-2, MU1377/9-2, GE2533/2-2]; This work was supported by Deutsche Forschungsgemeinschaft via, HE1964/13-2 to MH, MU1377/9-2 to YAM and GE2533/2-2 to RGG.
dc.language.isoen
dc.publisherACADEMIC PRESS INC ELSEVIER SCIENCE
dc.relation.ispartofPROTEIN EXPRESSION AND PURIFICATION
dc.subjectBiochemical Research Methods
dc.subjectBiochemistry & Molecular Biology
dc.subjectBiotechnology & Applied Microbiology
dc.subjectENTERICA
dc.subjectFRAGMENT
dc.subjectNON-FIMBRIAL ADHESIN
dc.subjectPATHOGENICITY
dc.subjectProtein expression
dc.subjectPurification
dc.subjectRECOGNITION
dc.subjectSalmonella
dc.subjectSECONDARY STRUCTURE PREDICTION
dc.subjectSEQUENCE
dc.subjectType I secretion system (T1SS)
dc.titleRecombinant protein production and purification of SiiD, SiiE and SiiF - Components of the SPI4-encoded type I secretion system from Salmonella Typhimurium
dc.typejournal article
dc.identifier.doi10.1016/j.pep.2020.105632
dc.identifier.isiISI:000531018500001
dc.description.volume172
dc.contributor.orcid0000-0003-0519-8928
dc.contributor.researcheridR-5397-2016
dc.identifier.eissn10960279
dc.publisher.place525 B ST, STE 1900, SAN DIEGO, CA 92101-4495 USA
dcterms.isPartOf.abbreviationProtein Expr. Purif.
dcterms.oaStatusGreen Submitted
crisitem.author.deptFB 05 - Biologie/Chemie-
crisitem.author.deptidfb05-
crisitem.author.orcid0000-0001-6604-6253-
crisitem.author.parentorgUniversität Osnabrück-
crisitem.author.netidHeMi480-
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