Four‐bond deuterium isotope effects on the chemical shifts of amide nitrogens in proteins

Abstract

An approach towards precision NMR measurements of four-bond deuterium isotope effects on the chemical shifts of backbone amide nitrogen nuclei in proteins is described. Three types of four-bond (15) N deuterium isotope effects are distinguished depending on the site of proton-to-deuterium substitution: (4)ΔN(N(i-1)D), (4)ΔN(N(i+1)D) and (4)ΔN(Cβ,(i-1)D). All the three types of isotope shifts are quantified in the (partially) deuterated protein ubiquitin. The (4)ΔN(N(i+1)D) and (4)ΔN(C(β,i-1)D) effects are by far the largest in magnitude and vary between 16 and 75 ppb and -18 and 46 ppb, respectively. A semi-quantitative correlation between experimental (4)ΔN(N(i+1)D) and (4)ΔN(C(β,i-1)D) values and the distances between nitrogen nuclei and the sites of (1)H-to-D substitution is noted. The largest isotope shifts in both cases correspond to the shortest inter-nuclear distances.

Isotope effects on chemical shifts in the study of hydrogen bonded biological systems

“…Strangely enough, Ile and Val had the opposite sign to the other residues. In relation to this, a more specific study of the effect of deuteriation at the C-β carbon has been conducted by Tugarinov [12]. Recently, Smith et al [13] found similar values for perdeuteriated proteins in a solid state study of HET fibrils, but they did not study these further for reasons of variying pH.…”