Protein (19)F NMR in Escherichia coli

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...

Full description

Saved in:
Bibliographic Details
Published in:Journal of the American Chemical Society 2010-01, Vol.132 (1), p.321-327
Main Authors: Li, Conggang, Wang, Gui-Fang, Wang, Yaqiang, Creager-Allen, Rachel, Lutz, Evan A, Scronce, Heidi, Slade, Kristin M, Ruf, Rebecca A S, Mehl, Ryan A, Pielak, Gary J
Format: Article
Language:English
Subjects:
Chymotrypsin - antagonists & inhibitors
Chymotrypsin - chemistry
Chymotrypsin - metabolism
Escherichia coli - chemistry
Halogenation
Molecular Weight
Nuclear Magnetic Resonance, Biomolecular
Protease Inhibitors - pharmacology
Proteins - chemistry
Proteins - genetics
Proteins - metabolism
Recombinant Proteins - chemistry
Recombinant Proteins - genetics
Recombinant Proteins - metabolism
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary: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.
ISSN:1520-5126
DOI:10.1021/ja907966n