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The Expanding Zoo of In-Cell Protein NMR
- Source :
- Biophysical journal. 115(9)
- Publication Year :
- 2018
-
Abstract
- In-cell NMR spectroscopy is a powerful tool to determine the properties of proteins and nucleic acids within living cells. In-cell NMR can give site-specific measurements of interactions, modifications, and dynamics as well as their modulation by the cellular environment. In-cell NMR requires selective incorporation of heavy isotopes into a protein of interest, either through the introduction of exogenously produced protein to a cell’s interior or the selective overexpression of a protein. We developed conditions to allow the use of Saccharomyces cerevisiae, which was chosen because of its genetic tractability, as a eukaryotic expression system for in-cell NMR. We demonstrate this technique using a fragment of S. cerevisiae Nsp1, an FG Nup. FG Nups are intrinsically disordered proteins containing phenylalanine (F)-glycine (G) repeats and form the selective barrier within the nuclear pore complex. Yeast FG Nups have previously been shown to be maintained in a highly dynamic state within living bacteria as measured by in-cell NMR. Interactions thought to stabilize this dynamic state are also present in the protein’s native organism, although site specificity of interaction is different between the two cytosols.
- Subjects :
- 0301 basic medicine
Cytoplasm
biology
Extramural
Chemistry
business.industry
Protein
Biophysics
010402 general chemistry
biology.organism_classification
01 natural sciences
Magnetic Resonance Imaging
0104 chemical sciences
Cell biology
03 medical and health sciences
Cytosol
030104 developmental biology
Text mining
Saccharomycetales
business
Hydrophobic and Hydrophilic Interactions
Subjects
Details
- ISSN :
- 15420086
- Volume :
- 115
- Issue :
- 9
- Database :
- OpenAIRE
- Journal :
- Biophysical journal
- Accession number :
- edsair.doi.dedup.....08428822c4bd02414f803d694bb6173a