title: Molecular-orientation analysis based on alignment-induced TROSY chemical shift changes.
abstract: We present a new NMR technique for determining the alignment tensor of a weakly aligned protein using only alignment-induced 15N transverse relaxation optimized spectroscopy (TROSY) chemical shift changes. Alignment-induced TROSY chemical shift changes reflect the combined contributions from two different anisotropic spin interactions including the residual dipolar couplings (RDCs) and the residual chemical shift anisotropy effects (RCSAs). We show here that these two residual anisotropic spin interactions' values, encoded in the TROSY chemical shift changes, can be used to determine a weakly aligned protein's alignment tensor. To prove the significance of this method, we show that our TROSY-based analysis gives the consistent alignment angles with those determined using RDCs for 15N-labeled ubiquitin (8.6 kDa) in an aligned medium, within an uncertainty range estimated by considering experimental and structural noises, being 5 degrees at most. Because our approach requires a pre-determined 15N CSA tensor value, we also estimated the uncertainties associated with the resultant alignment tensor values caused by variation in 15N CSA tensors. In spite of the significant variations in literature-reported 15N CSA tensors, they gave consistent orientation angles within an uncertainty range. These results ensure that our TROSY-based approach is a useful alternative to the RDC-based method to determine the alignment angles especially for large proteins in a weakly aligned state.
authors.affiliation: Department of structural biology, Biomolecular Engineering Research Institute, 6-2-3 Furuedai, Suita, Osaka 565-0874, Japan. tate@beri.or.jp
authors.forename: Shin-ichi
authors.initials: S
authors.lastname: Tate
authors.forename: Hideto
authors.initials: H
authors.lastname: Shimahara
authors.forename: Naoko
authors.initials: N
authors.lastname: Utsunomiya-Tate
mesh_terms: D016880:Anisotropy; D006801:Humans; D006860:Hydrogen Bonding; D009587:Nitrogen Isotopes; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods*; D011487:Protein Conformation; D025801:Ubiquitin / Q000737:chemistry*
chemical_list: D009587:Nitrogen Isotopes; D025801:Ubiquitin