Protein (19)F NMR in Escherichia coli. J Am Chem Soc 2010 Jan 13;132(1):321-7
Date
01/07/2010Pubmed ID
20050707Pubmed Central ID
PMC2815348DOI
10.1021/ja907966nScopus ID
2-s2.0-74849140345 (requires institutional sign-in at Scopus site) 211 CitationsAbstract
Although overexpression and (15)N enrichment facilitate the observation of resonances from disordered proteins in Escherichia coli, (15)N enrichment alone is insufficient for detecting most globular proteins. Here, we explain this dichotomy and overcome the problem while extending the capability of in-cell NMR by using (19)F-labeled proteins. Resonances from small (approximately 10 kDa) globular proteins containing the amino acid analogue 3-fluoro-tyrosine can be observed in cells, but for larger proteins the (19)F resonances are broadened beyond detection. Incorporating the amino acid analogue trifluoromethyl-L-phenylalanine allows larger proteins (up to 100 kDa) to be observed in cells. We also show that site-specific structural and dynamic information about both globular and disordered proteins can be obtained inside cells by using (19)F NMR.
Author List
Li C, Wang GF, Wang Y, Creager-Allen R, Lutz EA, Scronce H, Slade KM, Ruf RA, Mehl RA, Pielak GJAuthor
Yaqiang Wang PhD, MS Assistant Professor in the Biophysics department at Medical College of WisconsinMESH terms used to index this publication - Major topics in bold
ChymotrypsinEscherichia coli
Halogenation
Molecular Weight
Nuclear Magnetic Resonance, Biomolecular
Protease Inhibitors
Proteins
Recombinant Proteins
