caa a22 4500 445325194 CHVBK 20180317142718.0 cr unu---uuuuu 170323e20111101xx s 000 0 eng 10.1007/s10858-011-9559-4 doi (NATIONALLICENCE)springer-10.1007/s10858-011-9559-4 Automated sequence- and stereo-specific assignment of methyl-labeled proteins by paramagnetic relaxation and methyl-methyl nuclear overhauser enhancement spectroscopy [Elektronische Daten] [Vincenzo Venditti, Nicolas Fawzi, G. Clore] Methyl-transverse relaxation optimized spectroscopy is rapidly becoming the preferred NMR technique for probing structure and dynamics of very large proteins up to ~1MDa in molecular size. Data interpretation, however, necessitates assignment of methyl groups which still presents a very challenging and time-consuming process. Here we demonstrate that, in combination with a known 3D structure, paramagnetic relaxation enhancement (PRE), induced by nitroxide spin-labels incorporated at only a few surface-exposed engineered cysteines, provides fast, straightforward and robust access to methyl group resonance assignments, including stereoassignments for the methyl groups of leucine and valine. Neither prior assignments, including backbone assignments, for the protein, nor experiments that transfer magnetization between methyl groups and the protein backbone, are required. PRE-derived assignments are refined by 4D methyl-methyl nuclear Overhauser enhancement data, eliminating ambiguities and errors that may arise due to the high sensitivity of PREs to the potential presence of sparsely-populated transient states. Springer-Verlag, 2011 Paramagnetic relaxation enhancement nationallicence Methyl-TROSY nationallicence 1H-13C HMQC nationallicence Paramagnetic spin-labeling nationallicence Methyl group labeling nationallicence Venditti Vincenzo Laboratory of Chemical Physics, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, 20892-0520, Bethesda, MD, USA aut Fawzi Nicolas Laboratory of Chemical Physics, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, 20892-0520, Bethesda, MD, USA aut Clore G. Laboratory of Chemical Physics, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, 20892-0520, Bethesda, MD, USA aut Journal of Biomolecular NMR Springer Netherlands 51/3(2011-11-01), 319-328 0925-2738 51:3<319 2011 51 10858 https://doi.org/10.1007/s10858-011-9559-4 text/html Onlinezugriff via DOI 1 research-article jats NATIONALLICENCE 856 40 https://doi.org/10.1007/s10858-011-9559-4 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Venditti Vincenzo Laboratory of Chemical Physics, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, 20892-0520, Bethesda, MD, USA aut NATIONALLICENCE 700 1- Fawzi Nicolas Laboratory of Chemical Physics, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, 20892-0520, Bethesda, MD, USA aut NATIONALLICENCE 700 1- Clore G. Laboratory of Chemical Physics, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, 20892-0520, Bethesda, MD, USA aut NATIONALLICENCE 773 0- Journal of Biomolecular NMR Springer Netherlands 51/3(2011-11-01), 319-328 0925-2738 51:3<319 2011 51 10858 Metadata rights reserved Springer special CC-BY-NC licence nationallicence BK010053 XK010053 XK010000 NATIONALLICENCE NATIONALLICENCE NL-springer