Single molecule super-resolution imaging of proteins in living Salmonella enterica using self-labelling enzymes

Autor(en): Barlag, Britta
Beutel, Oliver
Janning, Dennis
Czarniak, Frederik
Richter, Christian P.
Kommnick, Carina
Goeser, Vera
Kurre, Rainer 
Fabiani, Florian
Erhardt, Marc
Piehler, Jacob 
Hensel, Michael 
Stichwörter: BACTERIAL-CELL BIOLOGY; CHEMOTAXIS; GENES; III SECRETION; LOCALIZATION; MICROCOMPARTMENTS; MICROSCOPY; Multidisciplinary Sciences; NON-FIMBRIAL ADHESIN; ORGANIZATION; Science & Technology - Other Topics; TRACKING
Erscheinungsdatum: 2016
Herausgeber: NATURE PUBLISHING GROUP
Journal: SCIENTIFIC REPORTS
Volumen: 6
Zusammenfassung: 
The investigation of the subcellular localization, dynamics and interaction of proteins and protein complexes in prokaryotes is complicated by the small size of the cells. Super-resolution microscopy ( SRM) comprise various new techniques that allow light microscopy with a resolution that can be up to ten-fold higher than conventional light microscopy. Application of SRM techniques to living prokaryotes demands the introduction of suitable fluorescent probes, usually by fusion of proteins of interest to fluorescent proteins with properties compatible to SRM. Here we describe an approach that is based on the genetically encoded self-labelling enzymes HaloTag and SNAP-tag. Proteins of interest are fused to HaloTag or SNAP-tag and cell permeable substrates can be labelled with various SRM-compatible fluorochromes. Fusions of the enzyme tags to subunits of a type I secretion system ( T1SS), a T3SS, the flagellar rotor and a transcription factor were generated and analysed in living Salmonella enterica. The new approach is versatile in tagging proteins of interest in bacterial cells and allows to determine the number, relative subcellular localization and dynamics of protein complexes in living cells.
ISSN: 20452322
DOI: 10.1038/srep31601

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