A Gateway-Based System for Fast Evaluation of Protein-Protein Interactions in Bacteria
DC Element | Wert | Sprache |
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dc.contributor.author | Wille, Thorsten | |
dc.contributor.author | Barlag, Britta | |
dc.contributor.author | Jakovljevic, Vladimir | |
dc.contributor.author | Hensel, Michael | |
dc.contributor.author | Sourjik, Victor | |
dc.contributor.author | Gerlach, Roman G. | |
dc.date.accessioned | 2021-12-23T16:21:46Z | - |
dc.date.available | 2021-12-23T16:21:46Z | - |
dc.date.issued | 2015 | |
dc.identifier.issn | 19326203 | |
dc.identifier.uri | https://osnascholar.ub.uni-osnabrueck.de/handle/unios/14013 | - |
dc.description.abstract | Protein-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. | |
dc.description.sponsorship | DFGGerman Research Foundation (DFG)European Commission [GE2533/1-1, SFB 944]; ERCEuropean Research Council (ERC)European Commission [294761]; This work was supported in part by the DFG through project P4 of the collaborative research centre SFB 944 `Physiology and dynamics of cellular microcompartments' at the University of Osnabruck (to M.H.) and GE2533/1-1 (to R.G.G.) and an ERC grant 294761 (to V.S.). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. | |
dc.language.iso | en | |
dc.publisher | PUBLIC LIBRARY SCIENCE | |
dc.relation.ispartof | PLOS ONE | |
dc.subject | 2-HYBRID SYSTEM | |
dc.subject | ENTERICA SEROVAR TYPHIMURIUM | |
dc.subject | ESCHERICHIA-COLI | |
dc.subject | FUSION PROTEINS | |
dc.subject | GAUSSIA LUCIFERASE | |
dc.subject | GENE-EXPRESSION | |
dc.subject | LIVING CELLS | |
dc.subject | Multidisciplinary Sciences | |
dc.subject | PCR PRODUCTS | |
dc.subject | SALMONELLA-ENTERICA | |
dc.subject | Science & Technology - Other Topics | |
dc.subject | SITE-SPECIFIC RECOMBINATION | |
dc.title | A Gateway-Based System for Fast Evaluation of Protein-Protein Interactions in Bacteria | |
dc.type | journal article | |
dc.identifier.doi | 10.1371/journal.pone.0123646 | |
dc.identifier.isi | ISI:000352588500102 | |
dc.description.volume | 10 | |
dc.description.issue | 4 | |
dc.contributor.orcid | 0000-0002-6082-4867 | |
dc.contributor.orcid | 0000-0001-6604-6253 | |
dc.publisher.place | 1160 BATTERY STREET, STE 100, SAN FRANCISCO, CA 94111 USA | |
dcterms.isPartOf.abbreviation | PLoS One | |
dcterms.oaStatus | Green Submitted, gold, Green Published | |
crisitem.author.dept | FB 05 - Biologie/Chemie | - |
crisitem.author.deptid | fb05 | - |
crisitem.author.orcid | 0000-0001-6604-6253 | - |
crisitem.author.parentorg | Universität Osnabrück | - |
crisitem.author.netid | HeMi480 | - |
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geprüft am 09.06.2024