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2015-04-09Zeitschriftenartikel DOI: 10.1371/journal.pone.0123646
A Gateway-Based System for Fast Evaluation of Protein-Protein Interactions in Bacteria
dc.contributor.authorWille, Thorsten
dc.contributor.authorBarlag, Britta
dc.contributor.authorJakovljevic, Vladimir
dc.contributor.authorHensel, Michael
dc.contributor.authorSourjik, Victor
dc.contributor.authorGerlach, Roman
dc.date.accessioned2018-05-07T18:12:50Z
dc.date.available2018-05-07T18:12:50Z
dc.date.created2015-04-21
dc.date.issued2015-04-09none
dc.identifier.otherhttp://edoc.rki.de/oa/articles/retmzJuAlBuY/PDF/24mlm8Tww3eYs.pdf
dc.identifier.urihttp://edoc.rki.de/176904/2052
dc.description.abstractProtein-protein interactions are important layers of regulation in all kingdoms of life. Identification and characterization of these interactions is one challenging task of the post-genomic era and crucial for understanding of molecular processes within a cell. Several methods have been successfully employed during the past decades to identify protein-protein interactions in bacteria, but most of them include tedious and time-consuming manipulations of DNA. In contrast, the MultiSite Gateway system is a fast tool for transfer of multiple DNA fragments between plasmids enabling simultaneous and site directed cloning of up to four fragments into one construct. Here we developed a new set of Gateway vectors including custom made entry vectors and modular Destination vectors for studying protein-protein interactions via Fluorescence Resonance Energy Transfer (FRET), Bacterial two Hybrid (B2H) and split Gaussia luciferase (Gluc), as well as for fusions with SNAP-tag and HaloTag for dual-color super-resolution microscopy. As proof of principle, we characterized the interaction between the Salmonella effector SipA and its chaperone InvB via split Gluc and B2H approach. The suitability for FRET analysis as well as functionality of fusions with SNAP- and HaloTag could be demonstrated by studying the transient interaction between chemotaxis response regulator CheY and its phosphatase CheZ.eng
dc.language.isoeng
dc.publisherRobert Koch-Institut
dc.subjectBacterial/geneticseng
dc.subjectEscherichia coli/geneticseng
dc.subjectBacterial Proteins/metabolismeng
dc.subjectMembrane Proteins/geneticseng
dc.subjectMembrane Proteins/metabolismeng
dc.subjectDNAeng
dc.subjectPlasmidseng
dc.subjectBacterial Proteins/geneticseng
dc.subjectSalmonella/geneticseng
dc.subjectEscherichia coli/metabolismeng
dc.subjectCloning Moleculareng
dc.subjectFluorescence Resonance Energy Transfereng
dc.subjectGenetic Vectorseng
dc.subjectLuciferases/geneticseng
dc.subjectMicrofilament Proteins/geneticseng
dc.subjectMicrofilament Proteins/metabolismeng
dc.subjectProtein Interaction Maps/geneticseng
dc.subjectSalmonella/metabolismeng
dc.subject.ddc610 Medizin
dc.titleA Gateway-Based System for Fast Evaluation of Protein-Protein Interactions in Bacteria
dc.typeperiodicalPart
dc.identifier.urnurn:nbn:de:0257-10039331
dc.identifier.doi10.1371/journal.pone.0123646
dc.identifier.doihttp://dx.doi.org/10.25646/1977
local.edoc.container-titlePLoS ONE
local.edoc.container-textWille T, Barlag B, Jakovljevic V, Hensel M, Sourjik V, et al. (2015) A Gateway-Based System for Fast Evaluation of Protein-Protein Interactions in Bacteria. PLoS ONE 10(4): e0123646.
local.edoc.fp-subtypeArtikel
local.edoc.type-nameZeitschriftenartikel
local.edoc.container-typeperiodical
local.edoc.container-type-nameZeitschrift
local.edoc.container-urlhttp://journals.plos.org/plosone/article?id=10.1371/journal.pone.0123646
local.edoc.container-publisher-namePublic Library of Science
local.edoc.container-volume10
local.edoc.container-issue4
local.edoc.container-year2015

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