A Gel-Encapsulated Bioreactor System for NMR Studies of Protein-Protein Interactions in Living Mammalian Cells

摘要

In-cell NMR spectroscopy is a method used to observe isotopically labeled molecules within living cells. The first in-cell NMR experiment was performed with an E. coli overexpressing a ~(15)N-labeled protein. For the first application of the in-cell NMR method with eukaryotic cells, isotopically labeled target proteins were introduced, by microinjection, into Xenopus laevis oocytes. Recently, Inomata et al. reported a novel in-cell NMR method utilizing a cell-penetrating tag, which opens the way for the application of in-cell NMR spectroscopy to mammalian cells. Our group also reported an in-cell NMR method for mammalian cells; we used a pore-forming toxin, streptolysin O (SLO), to introduce target proteins by diffusion. By using these methods, protein-drug interactions and intracellular post-translational modifications, such as phosphorylation and acetylation, were successfully detected in vivo.