Biomedical Engineering Reference
In-Depth Information
standard NMR spectrum of a protein, conferring to RDCs an important role
as motional probes.
8.4.1 Domain Dynamics
As in the structural case, the use of RDCs to study protein dynamics is most
intuitively appreciated when multiple RDCs are combined in a unit of known
structure for the study of domain motions within a larger macromolecular
ensemble. A first example considered diffusive motion of a-helices in magnetically
aligned cyanometmyoglobin, on the basis of RDCs and paramagnetic chemical
shifts.
33
This pioneering study laid the basis for numerous studies of domain-like
dynamic behavior using similar approaches, for proteins and RNA or
oligosaccarides.
34-40
The basis for such analyses is that the effective alignment
tensor elements shown in eqn (8.2) for each domain will be dependent not onlyon
the orientation of the domain within the molecular frame, but will also be
influenced by the differential domain dynamics. Comparison of the effective
alignment tensors of different domains can then report on the direction and
amplitude of intra-domain motions. It is important to note however, that this kind
of analysis is ultimately limited by the absence of an absolute external reference
frame: re-orientational motions of two domains that are of equal size are virtually
impossible to detect in this way as the alignment tensor parameters of both
domains would be essentially identical. For this reason RDCs have been
combined with paramagnetic pseudo-contact shifts, induced by the presence of
a native or non-native paramagnetic probe attached to one of the two
domains.
33,41-45
These effects are dependent both on the angular and radial
terms that are dependent on the distance between the electron spin and the nuclear
spins, and therefore carry a greater potential for studying domain motions.
8.4.2 Local Backbone Dynamics
Because RDCs are sensitive to a population-weighted average of all
conformations sampled up to the millisecond, they are exquisitely sensitive
to the structural details of local conformational dynamics and this aspectwill
be the subject of the remainder of this chapter (Figure 8.2). The straightfor-
ward averaging properties of RDCs allows for rigorous interpretation in terms
of local dynamic modes and amplitudes. The different assumptions that are
made when analysing protein dynamics from RDCs, and the validity of the
resulting dynamic description, can be statistically tested using standard
statistical tests and cross-validation, making RDCs potentially even more
powerful than classical spin-relaxation experiments, that report on dynamic
timescales on the in the pico- to nanosecond timescale. The disadvantage of
RDCs is that they report on motions occurring on all timescales up to the
millisecond, and that it is not possible, from the data alone, to distinguish
between these different timescales. Comparison with dynamic amplitudes
extracted from RDCs and spin-relaxation, measured on the same dipolar
