Cell-free protein synthesis for analysis by NMR spectroscopy.
Cell-free protein synthesis offers fast and inexpensive access to selectively isotope labeled proteins that can be measured by NMR spectroscopy in the presence of all the unlabeled proteins in the reaction mixture. No chromatographic purification is required. Using an extract from Escherichia coli in a simple dialysis system, the target protein can be prepared at a typical concentration of about 1 mg/ml, which is sufficient for subsequent analysis by NMR. This chapter describes in detail the protocol used in the authors' laboratory.
Research School of Chemistry, Australian National University, Canberra, Australia.
Margit A
MA
Apponyi
Kiyoshi
K
Ozawa
Nicholas E
NE
Dixon
Gottfried
G
Otting
D002474:Cell-Free System; D004926:Escherichia coli / Q000737:chemistry; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D007553:Isotope Labeling; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D009587:Nitrogen Isotopes; D014176:Protein Biosynthesis
D029968:Escherichia coli Proteins; D009587:Nitrogen Isotopes