Biomedical Engineering Reference
In-Depth Information
agreement with a static description, and encouraged the use of RDCs for
structure refinement.
RDCs are now routinely incorporated into NMR structure-determination
protocols,
19-22
most often in combination with a full set of internuclear
distance restraints derived from NOESY experiments.
23
Not surprisingly this
combination leads to more precise structural bundles, as the conformational
space is necessarily more restricted when more constraints are applied. In
general, both accuracy and precision can be expected to improve, although it is
important to exercise some caution as to the inherent structural information
that is available when using RDCs as constraints. As shown in Figure 8.1, a
single RDC has a very broad angular degeneracy, such that refinement using
only RDCs from
15
N-
1
H
N
pairs in a protein carries little additional structural
information on its own. The combination of multiple RDCs present in a
structural element whose three-dimensional structure is known can overcome
this degeneracy, so that the orientation of this element has only a four
equivalent orientations.
24
This characteristic allows for the determination of
the relative orientation of structural domains, which is possibly the most
intuitively novel application of liquid-crystal NMR to protein structure: for
the first time, it was possible to measure structural information from distant
regions of the same macromolecule or macromolecular complex that could be
used to determine the relative orientation of the component parts relativeto
each other.
25
In combination with chemical shift perturbations, or inter-
molecular distance constraints, RDCs have thus been successfully appliedto
the study of large supramolecular complexes that were previously inaccessible
to NMR studies.
24-29
On a more local level, it has recently been demonstrated
that a peptide plane, even with the measurement of multiple RDCs that are
arranged in different directions, still has 16 equally valid orientations with
respect to the molecular frame.
30
These degeneracies can be further overcome
by combining data from differently aligning media.
24
The intrinsic structural
information then becomes far more powerful, even leading to the determina-
tion of the three-dimensional protein structure using only RDCs.
31,32
8.4 Residual Dipolar Couplings for the Study of Protein
Dynamics
As mentioned above, RDCs also provide information about time- and
ensemble-averaged conformational processes occurring on timescales up to
milliseconds, and thereby contain information for understanding biomolecular
motions that is unavailable from other sources. All measured first-order
interactions are modulated by motion occurring on timescales that are faster
than the magnitude of the interaction. Experimental RDCs are measured in an
absolute range of up to tens of Hz, corresponding to dynamic correlation times
of a few tens of milliseconds, and broadly coinciding with the chemical shift
coalescence limit. RDCs are therefore sensitive to all timescales measured in a
