Abstract
NMR spectroscopy enables structural analyses of proteins and has been widely used in the structural biology field in recent decades. NMR spectroscopy can be applied to proteins inside living cells, allowing characterization of their structures and dynamics in intracellular environments. The simplest "in-cell NMR" approach employs bacterial cells; in this approach, live Escherichia coli cells overexpressing a specific protein are subjected to NMR. The cells are grown in an NMR active isotope-enriched medium to ensure that the overexpressed proteins are labeled with the stable isotopes. Thus the obtained NMR spectra, which are derived from labeled proteins, contain atomic-level information about the structure and dynamics of the proteins. Recent progress enables us to work with higher eukaryotic cells such as HeLa and HEK293 cells, for which a number of techniques have been developed to achieve isotope labeling of the specific target protein. In this review, we describe successful use of electroporation for in-cell NMR. In addition, (19)F-NMR to characterize protein-ligand interactions in cells is presented. Because (19)F nuclei rarely exist in natural cells, when (19)F-labeled proteins are delivered into cells and (19)F-NMR sign...Continue Reading
References
Jan 25, 2006·Nature Methods·David S BurzAlexander Shekhtman
Oct 13, 2006·Journal of Biomolecular NMR·Tomomi SakaiMasahiro Shirakawa
Apr 5, 2007·Nature Protocols·David S BurzAlexander Shekhtman
Apr 5, 2007·Nature Protocols·Zach SerberVolker Dötsch
Feb 26, 2008·Nature Structural & Molecular Biology·Philipp SelenkoGerhard Wagner
Jun 25, 2008·Annual Review of Biophysics·Huan-Xiang ZhouAllen P Minton
Mar 6, 2009·Nature·Daisuke SakakibaraYutaka Ito
Mar 6, 2009·Nature·Kohsuke InomataMasahiro Shirakawa
Mar 31, 2009·Nature Chemical Biology·Shinya TsukijiItaru Hamachi
May 9, 2009·Journal of Medicinal Chemistry·Jingjing XieAlexander Shekhtman
Jul 17, 2009·Journal of the American Chemical Society·Shinji OginoIchio Shimada
May 4, 2011·Journal of the American Chemical Society·Alexander P SchlesingerGary J Pielak
Jul 5, 2012·Journal of the American Chemical Society·Karl BertrandAlexander Shekhtman
Oct 23, 2012·Nature·Sang Ho ParkStanley J Opella
Dec 1, 2012·Angewandte Chemie·Satoshi KuboIchio Shimada
Feb 2, 2013·Science·H J MaminD Rugar
Feb 2, 2013·Science·T StaudacherJ Wrachtrup
Feb 27, 2013·Chemical Communications : Chem Comm·Yousuke TakaokaItaru Hamachi
Mar 5, 2013·Nature Chemical Biology·Lucia BanciA Radu Aricescu
Jun 26, 2013·Chemistry & Biology·Lucia BanciErica Secci
Jul 4, 2013·Journal of the American Chemical Society·Jens DanielssonMikael Oliveberg
Aug 24, 2013·Journal of the American Chemical Society·Laura M LuhVolker Dötsch
Aug 31, 2013·PloS One·Christopher A WaudbyJohn Christodoulou
Nov 13, 2013·Proceedings of the National Academy of Sciences of the United States of America·Mohona SarkarGary J Pielak
Jun 28, 2014·Biochimica Et Biophysica Acta·Letizia BarbieriLucia Banci