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