Dataset Information

Mapping protein-specific micro-environments in live cells by fluorescence lifetime imaging of a hybrid genetic-chemical molecular rotor tag.

ABSTRACT: The micro-viscosity and molecular crowding experienced by specific proteins can regulate their dynamics and function within live cells. Taking advantage of the emerging TMP-tag technology, we present the design, synthesis and application of a hybrid genetic-chemical molecular rotor probe whose fluorescence lifetime can report protein-specific micro-environments in live cells.

SUBMITTER: Gatzogiannis E

PROVIDER: S-EPMC3775488 | biostudies-literature | 2012 Sep

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Mapping protein-specific micro-environments in live cells by fluorescence lifetime imaging of a hybrid genetic-chemical molecular rotor tag.

Gatzogiannis Evangelos E Chen Zhixing Z Wei Lu L Wombacher Richard R Kao Ya-Ting YT Yefremov Grygorii G Cornish Virginia W VW Min Wei W

Chemical communications (Cambridge, England) 20120724 69

The micro-viscosity and molecular crowding experienced by specific proteins can regulate their dynamics and function within live cells. Taking advantage of the emerging TMP-tag technology, we present the design, synthesis and application of a hybrid genetic-chemical molecular rotor probe whose fluorescence lifetime can report protein-specific micro-environments in live cells. ...[more]

PMID: 22825488

Similar Datasets

Project description:Live-cell fluorescence imaging of methanogenic archaea has been limited due to the strictly anoxic conditions required for growth and issues with autofluorescence associated with electron carriers in central metabolism. Here, we show that the fluorescence-activating and absorption-shifting tag (FAST) complexed with the fluorogenic ligand 4-hydroxy-3-methylbenzylidene-rhodanine (HMBR) overcomes these issues and displays robust fluorescence in Methanococcus maripaludis. We also describe a mechanism to visualize cells under anoxic conditions using a fluorescence microscope. Derivatives of FAST were successfully applied for protein abundance analysis, subcellular localization analysis, and determination of protein-protein interactions. FAST fusions to both formate dehydrogenase (Fdh) and F420-reducing hydrogenase (Fru) displayed increased fluorescence in cells grown on formate-containing medium, consistent with previous studies suggesting the increased abundance of these proteins in the absence of H2. Additionally, FAST fusions to both Fru and the ATPase associated with the archaellum (FlaI) showed a membrane localization in single cells observed using anoxic fluorescence microscopy. Finally, a split reporter translationally fused to the alpha and beta subunits of Fdh reconstituted a functionally fluorescent molecule in vivo via bimolecular fluorescence complementation. Together, these observations demonstrate the utility of FAST as a tool for studying members of the methanogenic archaea. IMPORTANCE Methanogenic archaea are important members of anaerobic microbial communities where they catalyze essential reactions in the degradation of organic matter. Developing additional tools for studying the cell biology of these organisms is essential to understanding them at a mechanistic level. Here, we show that FAST, in combination with the fluorogenic ligand HMBR, can be used to monitor protein dynamics in live cells of M. maripaludis. The application of FAST holds promise for future studies focused on the metabolism and physiology of methanogenic archaea.

Dataset Information

Mapping protein-specific micro-environments in live cells by fluorescence lifetime imaging of a hybrid genetic-chemical molecular rotor tag.

Publications

Mapping protein-specific micro-environments in live cells by fluorescence lifetime imaging of a hybrid genetic-chemical molecular rotor tag.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets