In-cell NMR of intrinsically disordered proteins in prokaryotic cells

In-cell NMR, i.e., the acquisition of heteronuclear multidimensional NMR of biomacromolecules inside living cells, is, to our knowledge, the only method for investigating the three-dimensional structure and dynamics of proteins at atomic detail in the intracellular environment. Since the inception of the method, intrinsically disordered proteins have been regarded as particular targets for in-cell NMR, due to their expected sensitivity to the molecular crowding in the intracellular environment. While both prokaryotic and eukaryotic cells can be used as host cells for in-cell NMR, prokaryotic in-cell NMR, particularly employing commonly used protein overexpression systems in Escherichia coli cells, is the most accessible approach. In this chapter we describe general procedures for obtaining in-cell NMR spectra in E. coli cells.