Application of H(N)CA,CO-E.COSY experiments for calibrating the φ angular dependences of vicinal couplings J(C′i−1,Hiα), J(C′i−1,Ciβ) and J(C′i−1,C′i) in proteins

摘要: A triple-resonance NMR technique suitable for the determination ofcarbonyl-related couplings in polypeptide systems is introduced. Theapplication of three novel pulse sequences to uniformly13C/15N-enriched proteins yields E.COSY-likemultiplet patterns exhibiting either one the3J(C′i−1,Hiα), 3J(C′i−1,Ciβ) and3J(C′i−1,C′i)coupling constants indirectly detected 13C′dimension, depending on passive spin selected. The experiments aredemonstrated with oxidized flavodoxin from Desulfovibrio vulgaris. On thebasis J-values measured and backbone φ-angles derived ahigh-resolution X-ray structure protein, associated Karplusequations were reparametrized. root-mean-square differences between theexperimental coupling those predicted by optimized Karpluscurves are 0.41, 0.33 0.32 Hz for3J(C′i−1,Hiα),3J(C′i−1,Ciβ) and3J(C′i−1,C′i),respectively. results compared Karplus parameters previouslypublished same couplings.