Breaking symmetry in the structure determination of (large) symmetric protein dimers.
We demonstrate a novel methodology to disrupt the symmetry in the NMR spectra of homodimers. A paramagnetic probe is introduced sub-stoichiometrically to create an asymmetric system with the paramagnetic probe residing on only one monomer within the dimer. This creates sufficient magnetic anisotropy for resolution of symmetry-related overlapped resonances and, consequently, detection of pseudocontact shifts and residual dipolar couplings specific to each monomeric component. These pseudocontact shifts can be readily incorporated into existing structure refinement calculations and enable determination of monomer orientation within the dimeric protein. This methodology can be widely used for solution structure determination of symmetric dimers.
Gaponenko
Vadim
V
Structural Biophysics Laboratory, National Cancer Institute, PO Box B, Frederick, MD 21702-1201, USA.
Altieri
Amanda S
AS
Li
Jess
J
Byrd
R Andrew
RA
D004268:DNA-Binding Proteins / Q000737:chemistry; D019281:Dimerization; D004492:Edetic Acid / Q000737:chemistry; D046911:Macromolecular Substances; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000737:chemistry*; D050797:STAT4 Transcription Factor; D015534:Trans-Activators / Q000737:chemistry
D004268:DNA-Binding Proteins; D046911:Macromolecular Substances; D011506:Proteins; D050797:STAT4 Transcription Factor; D015534:Trans-Activators; D004492:Edetic Acid