A dipolar coupling based strategy for simultaneous resonance assignment and structure determination of protein backbones.
A new approach for simultaneous protein backbone resonance assignment and structure determination by NMR is introduced. This approach relies on recent advances in high-resolution NMR spectroscopy that allow observation of anisotropic interactions, such as dipolar couplings, from proteins partially aligned in field ordered media. Residual dipolar couplings are used for both geometric information and a filter in the assembly of residues in a sequential manner. Experimental data were collected in less than one week on a small redox protein, rubredoxin, that was 15N enriched but not enriched above 1% natural abundance in 13C. Given the acceleration possible with partial 13C enrichment, the protocol described should provide a very rapid route to protein structure determination. This is critical for the structural genomics initiative where protein expression and structural determination in a high-throughput manner will be needed.
Tian
F
F
Southeast Collaboratory for Structural Genomics, University of Georgia, Athens, Georgia 30602-4712, USA.
Valafar
H
H
Prestegard
J H
JH
D008958:Models, Molecular; D009587:Nitrogen Isotopes; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods*; D011487:Protein Conformation; D020027:Pyrococcus furiosus / Q000737:chemistry; D012416:Rubredoxins / Q000737:chemistry*
D009587:Nitrogen Isotopes; D012416:Rubredoxins