Attenuated T2 relaxation by mutual cancellation of
dipole-dipole coupling and chemical shift anisotropy indicates
an avenue to NMR structures of very large biological
macromolecules in solution.

Pervushin K, Riek R, Wider G, Wuthrich K

Institut fur Molekularbiologie und Biophysik Eidgenossische Technische Hochschule Honggerberg
CH-8093 Zurich, Switzerland.

Fast transverse relaxation of 1H, 15N, and 13C by dipole-dipole coupling (DD) and chemical shift
anisotropy (CSA) modulated by rotational molecular motions has a dominant impact on the size limit
for biomacromolecular structures that can be studied by NMR spectroscopy in solution. Transverse
relaxation-optimized spectroscopy (TROSY) is an approach for suppression of transverse relaxation in
multidimensional NMR experiments, which is based on constructive use of interference between DD
coupling and CSA. For example, a TROSY-type two-dimensional 1H,15N-correlation experiment
with a uniformly 15N-labeled protein in a DNA complex of molecular mass 17 kDa at a 1H frequency
of 750 MHz showed that 15N relaxation during 15N chemical shift evolution and 1HN relaxation
during signal acquisition both are significantly reduced by mutual compensation of the DD and CSA
interactions. The reduction of the linewidths when compared with a conventional two-dimensional
1H,15N-correlation experiment was 60% and 40%, respectively, and the residual linewidths were 5 Hz
for 15N and 15 Hz for 1HN at 4 degrees C. Because the ratio of the DD and CSA relaxation rates is
nearly independent of the molecular size, a similar percentagewise reduction of the overall transverse
relaxation rates is expected for larger proteins. For a 15N-labeled protein of 150 kDa at 750 MHz and
20 degrees C one predicts residual linewidths of 10 Hz for 15N and 45 Hz for 1HN, and for the
corresponding uniformly 15N,2H-labeled protein the residual linewidths are predicted to be smaller
than 5 Hz and 15 Hz, respectively. The TROSY principle should benefit a variety of multidimensional
solution NMR experiments, especially with future use of yet somewhat higher polarizing magnetic
fields than are presently available, and thus largely eliminate one of the key factors that limit work
with larger molecules.