title: Automated NMR structure determination of stereo-array isotope labeled ubiquitin from minimal sets of spectra using the SAIL-FLYA system.
abstract: Stereo-array isotope labeling (SAIL) has been combined with the fully automated NMR structure determination algorithm FLYA to determine the three-dimensional structure of the protein ubiquitin from different sets of input NMR spectra. SAIL provides a complete stereo- and regio-specific pattern of stable isotopes that results in sharper resonance lines and reduced signal overlap, without information loss. Here we show that as a result of the superior quality of the SAIL NMR spectra, reliable, fully automated analyses of the NMR spectra and structure calculations are possible using fewer input spectra than with conventional uniformly 13C/15N-labeled proteins. FLYA calculations with SAIL ubiquitin, using a single three-dimensional "through-bond" spectrum (and 2D HSQC spectra) in addition to the 13C-edited and 15N-edited NOESY spectra for conformational restraints, yielded structures with an accuracy of 0.83-1.15 A for the backbone RMSD to the conventionally determined solution structure of SAIL ubiquitin. NMR structures can thus be determined almost exclusively from the NOESY spectra that yield the conformational restraints, without the need to record many spectra only for determining intermediate, auxiliary data of the chemical shift assignments. The FLYA calculations for this report resulted in 252 ubiquitin structure bundles, obtained with different input data but identical structure calculation and refinement methods. These structures cover the entire range from highly accurate structures to seriously, but not trivially, wrong structures, and thus constitute a valuable database for the substantiation of structure validation methods.
authors: Institute of Biophysical Chemistry, Center for Biomolecular Magnetic Resonance, Goethe University Frankfurt am Main, Max-von-Laue-Str. 9, 60438 Frankfurt am Main, Germany.
authors: Teppei
authors: T
authors: Ikeya
authors: Mitsuhiro
authors: M
authors: Takeda
authors: Hitoshi
authors: H
authors: Yoshida
authors: Tsutomu
authors: T
authors: Terauchi
authors: Jun-Goo
authors: JG
authors: Jee
authors: Masatsune
authors: M
authors: Kainosho
authors: Peter
authors: P
authors: Güntert
mesh_terms: D000465:Algorithms; D002247:Carbon Isotopes / Q000737:chemistry; D007553:Isotope Labeling; D008958:Models, Molecular; D009587:Nitrogen Isotopes / Q000737:chemistry; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods*; D015203:Reproducibility of Results; D025801:Ubiquitin / Q000737:chemistry*
chemical_list: D002247:Carbon Isotopes; D009587:Nitrogen Isotopes; D025801:Ubiquitin