title: One-sample approach to determine the relative orientations of proteins in ternary and binary complexes from residual dipolar coupling measurements.
abstract: We present a procedure that supports the acquisition of (1)H-(15)N residual dipolar coupling (RDC) values for individual subunits in binary or ternary protein assemblies from a single experimental sample. Our method relies on asymmetric labeling of each subunit with the following scheme: species A uniformly with (15)N, species B uniformly with (15)N and (13)C, and species C uniformly with (15)N but selectively with (13)C' or (13)C(alpha). Because only a single sample is required, the approach obviates the need for preparing multiple samples and eliminates potential errors introduced from differences in sample conditions. Because numerous biological processes rely on protein assemblies or transient interactions, this method should be well suited for a wide range of future applications.
authors.affiliation: National Magnetic Resonance Facility at Madison, Department of Biochemistry, University of Wisconsin-Madison, Madison, Wisconsin 53706-1544, USA.
authors.forename: Marco
authors.initials: M
authors.lastname: Tonelli
authors.forename: Larry R
authors.initials: LR
authors.lastname: Masterson
authors.forename: Gabriel
authors.initials: G
authors.lastname: Cornilescu
authors.forename: John L
authors.initials: JL
authors.lastname: Markley
authors.forename: Gianluigi
authors.initials: G
authors.lastname: Veglia
mesh_terms: D002247:Carbon Isotopes; D009587:Nitrogen Isotopes; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods*; D011487:Protein Conformation; D011506:Proteins / Q000737:chemistry*
chemical_list: D002247:Carbon Isotopes; D009587:Nitrogen Isotopes; D011506:Proteins