Field Strength Dependence ofTROSY for 15N1H Groups

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For the amide proton of a 15N-1H moiety, the two interfering relaxation mechanisms used in TROSY are DD relaxation between the proton and nitrogen spins and CSA relaxation of the proton. In general, DD and CSA relaxation are not equal in magnitude and cannot compensate each other. However, the DD interaction does not depend on the strength of the static magnetic field, whereas the CSA relaxation increases with larger magnetic fields, tte optimal TROSY effect for one doublet component can thus be obtained by choosing the appropriate field strength, where its relaxation rate will be near zero. For amide protons in polypeptides, this "magic field" is about 23.5 T, corresponding to a proton resonance frequency of approximately 1,000 MHz. tte 15N nucleus in an amide moiety shows a similar interference between 15N-1H DD interaction and its CSA, with a minimal transverse relaxation rate at magnetic field strengths corresponding to a proton resonance frequency of approximately 900 MHz, surprisingly very close to the value for the amide protons.

In practice, some deviations are expected from the "magic field" calculated for an isolated two-spin system, since the CSA varies slightly for different amide moieties. Further, residual DD coupling of amide group spins (especially of amide protons) with remote protons gives rise to relaxation that cannot be compensated by the TROSY effect, tte residual relaxation can be minimized in uniformly deuterated proteins, for which remote DD couplings are limited to nearby amide protons. In general, one approaches the optimal TROSY effect for peptide 15N-1H groups, manifested in optimal resolution and sensitivity of NMR spectra, at the highest presently available XH frequencies of 900 MHz (Pervushin et al. 1997) using perdeuterated samples in aqueous solutions.

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