Biomedical Engineering Reference
In-Depth Information
4.3.2.2
Disadvantages
Without
knowledge
of
an
alignment
tensor
RDCs
are
not
immediately
structurally instructive.
65,66
Only if a good approximate model of the overall
fitted
67
predicted
68
structure
exists
already,
can
alignment
tensors
be
or
accurately.
Spurious errors and distortions might stem from interaction with the
alignment media
69
or neglection of dynamics.
70
Experimentally, it is sometimes
difficult to find suitable alignment conditions and some trial and error is
required. Protein sample might be recoverable after it has been mixed with
alignment media.
4.3.2.3 The Bottom Line
RDCs could potentially be the workhorse of NMR structure determination for
high molecular weight structures.
63
For de novo structure determination,
however, dedicated sampling methods are required that can find the global
minimum in the non-instructive and rugged RDC restraint energy landscape
(Section 4.4).
4.3.3 Paramagnetic Relaxation Enhancement
PRE is caused by magnetic dipolar interactions between a nucleus and the
unpaired electrons of a paramagnetic center, and results in an increase in the
relaxation rate of the nuclear magnetisation.
71
The magnitude of the PRE is
proportional to r
26
(where r is the distance between the nucleus of interest and
the paramagnetic centre). Owing to the large magnetic moment of an unpaired
electron, the effect is detectable for sizeable separations (up to 34
˚
for Mn
2+
).
1
PREs are popular for elucidating complex structures. In contrast to NOEs,
PREs can provide restraints across sizeable separations and thus readily
capture the relative positions of the domains. The most useful spin-labels for
PRE measurements are ones that have an unpaired electron with an isotropic
g-tensor.
72
Examples include nitroxide spin-labels and EDTA-Mn
2+
. The
isotropic g-tensor ensures that the paramagnetic centre does not give rise to
pseudo-contact shifts (see below).
72
If conformational exchange dynamics on the millisecond timescale or faster
are present, the PRE is dominated by the species with shorter distances. This
can be exploited to resolve fast dynamic processes (Chapter 1), or to obtain
structures of weak-binding complexes,
3
or—if neglected—can lead to
spuriously short restraints. If the paramagnetic centre is attached via a
molecular linker it is important to explicitly model the motion of the tag by
ensemble approaches.
73
An interesting alternative to PRE measurements with protein-bound
paramagnetic centres is the introduction of paramagnetic centres bound to
