In-cell protein NMR and protein leakage.
In-cell nuclear magnetic resonance spectroscopy is a tool for studying proteins under physiologically relevant conditions. In some instances, however, protein signals from leaked protein are observed in the liquid surrounding the cells. Here, we examine the expression of four proteins in Escherichia coli. We describe the controls that should be used for in-cell NMR experiments and show that leakage is likely when the protein being studied exceeds ∼20% of the total cellular protein.
Department of Chemistry, University of North Carolina at Chapel Hill, North Carolina 27599, USA.
Christopher O
CO
Barnes
Gary J
GJ
Pielak
D004926:Escherichia coli / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000096:biosynthesis* / Q000737:chemistry; D006801:Humans; D019906:Nuclear Magnetic Resonance, Biomolecular; D010455:Peptides / Q000737:chemistry; D010940:Plant Proteins / Q000096:biosynthesis* / Q000737:chemistry; D014176:Protein Biosynthesis; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000737:chemistry; D025801:Ubiquitin / Q000096:biosynthesis* / Q000737:chemistry; D051844:alpha-Synuclein / Q000096:biosynthesis* / Q000737:chemistry
D029968:Escherichia coli Proteins; D010455:Peptides; D010940:Plant Proteins; D011994:Recombinant Proteins; D025801:Ubiquitin; D051844:alpha-Synuclein; C093336:chymotrypsin inhibitor 2; C084501:hdeA protein, E coli