title: Improving the accuracy of NMR structures of large proteins using pseudocontact shifts as long-range restraints.
abstract: We demonstrate improved accuracy in protein structure determination for large (>/=30 kDa), deuterated proteins (e.g. STAT4(NT)) via the combination of pseudocontact shifts for amide and methyl protons with the available NOEs in methyl-protonated proteins. The improved accuracy is cross validated by Q-factors determined from residual dipolar couplings measured as a result of magnetic susceptibility alignment. The paramagnet is introduced via binding to thiol-reactive EDTA, and multiple sites can be serially engineered to obtain data from alternative orientations of the paramagnetic anisotropic susceptibility tensor. The technique is advantageous for systems where the target protein has strong interactions with known alignment media.
authors.affiliation: Structural Biophysics Laboratory, National Cancer Institute, P.O. Box B, Frederick, MD 21702-1201, USA.
authors.forename: Vadim
authors.initials: V
authors.lastname: Gaponenko
authors.forename: Siddhartha P
authors.initials: SP
authors.lastname: Sarma
authors.forename: Amanda S
authors.initials: AS
authors.lastname: Altieri
authors.forename: David A
authors.initials: DA
authors.lastname: Horita
authors.forename: Jess
authors.initials: J
authors.lastname: Li
authors.forename: R Andrew
authors.initials: RA
authors.lastname: Byrd
mesh_terms: D000818:Animals; D004268:DNA-Binding Proteins / Q000737:chemistry; D008958:Models, Molecular; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods*; D010455:Peptides / Q000737:chemistry; D011506:Proteins / Q000737:chemistry*; D015203:Reproducibility of Results; D050797:STAT4 Transcription Factor; D015534:Trans-Activators / Q000737:chemistry
chemical_list: D004268:DNA-Binding Proteins; D010455:Peptides; D011506:Proteins; D050797:STAT4 Transcription Factor; D015534:Trans-Activators