An Empirical Correlation between Amide Deuterium Isotope Effects on 13Cα Chemical Shifts and Protein Backbone Conformation

Three-dimensional, triple resonance NMR techniques are described for measurement of two-bond (intraresidual) and three-bond (sequential) amide deuterium isotope effects on 13Cα chemical shifts. Measurements were carried out for uniformly 15N and 13C labeled human ubiquitin equilibrated in a 50% H2O/50% D2O mixed solvent. The three-bond isotope shift, 3ΔCα(ND), ranges from about 10−50 ppb, and, except for residues with positive φ angles and residues followed by Gly, its magnitude is described by 3ΔCα(ND) = 30.1 + 22.2 sin(ψ) ± 3.4 ppb. The two-bond isotope shift, 2ΔCα(ND), ranges from 70 to 116 ppb and is also dominated by the local backbone geometry at the Cα position:  2ΔCα(ND) = 93.1 + 10.1 sin(φ+62°) + 12.0 sin(ψ+42°) ± 4.1 ppb.