title: 15N and13C- SOFAST-HMQC editing enhances 3D-NOESY sensitivity in highly deuterated, selectively [1H,13C]-labeled proteins.
abstract: The ongoing NMR method development effort strives for high quality multidimensional data with reduced collection time. Here, we apply 'SOFAST-HMQC' to frequency editing in 3D NOESY experiments and demonstrate the sensitivity benefits using highly deuterated and15N, methyl labeled samples in H2O. The experiments benefit from a combination of selective T1relaxation (or L-optimized effect), from Ernst angle optimization and, in certain types of experiments, from using the mixing time for both NOE buildup and magnetization recovery. This effect enhances sensitivity by up to 2.4× at fast pulsing versus reference HMQC sequences of same overall length and water suppression characteristics. Representative experiments designed to address interesting protein NMR challenges are detailed. Editing capabilities are exploited with heteronuclear15N,13C-edited, or with diagonal-free13C aromatic/methyl-resolved 3D-SOFAST-HMQC-NOESY-HMQC. The latter experiment is used here to elucidate the methyl-aromatic NOE network in the hydrophobic core of the 19 kDa FliT-FliJ flagellar protein complex. Incorporation of fast pulsing to reference experiments such as 3D-NOESY-HMQC boosts digital resolution, simplifies the process of NOE assignment and helps to automate protein structure determination.
authors.affiliation: Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minneapolis, MN, 55455, USA. rossip@umn.edu.
authors.forename: Paolo
authors.identifier: 0000-0002-6041-0664
authors.initials: P
authors.lastname: Rossi
authors.affiliation: Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minneapolis, MN, 55455, USA.
authors.forename: Youlin
authors.initials: Y
authors.lastname: Xia
authors.affiliation: Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minneapolis, MN, 55455, USA.
authors.forename: Nandish
authors.initials: N
authors.lastname: Khanra
authors.affiliation: Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minneapolis, MN, 55455, USA. vegli001@umn.edu.
authors.forename: Gianluigi
authors.initials: G
authors.lastname: Veglia
authors.affiliation: Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minneapolis, MN, 55455, USA. ckalodim@umn.edu.
authors.forename: Charalampos G
authors.initials: CG
authors.lastname: Kalodimos
mesh_terms: D000465:Algorithms; D003903:Deuterium; D007553:Isotope Labeling; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D008962:Models, Theoretical; D009587:Nitrogen Isotopes; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods; D011506:Proteins / Q000737:chemistry*; D011522:Protons
chemical_list: D009587:Nitrogen Isotopes; D011506:Proteins; D011522:Protons; D003903:Deuterium
keywords: Fast pulsing data collection; Liquid state NMR; NOESY; SOFAST; SOFAST–NOESY