Chemistry Reference
In-Depth Information
[
76
]. Detection of exchange crosspeaks in
1
H spectra is often difficult in large
proteins because of interference from the many
1
H NOE cross peaks. In contrast,
observation of such exchange through
15
N spectroscopy has the advantage that only
exchange peaks are observed. The
15
N Z-exchange two-dimensional spectra are
acquired using a pulse sequence similar to
15
N
R
1
experiment but with
t
1
chemical
shift evolution period prior to the Z-mixing (relaxation) period [
4
,
77
-
79
]. The
measured rates of
exchange provide
information useful
to characterize
ligand-protein and protein-protein interactions.
4 Relaxation Dispersion Experiments
In contrast to
15
N
R
1
,
R
2
, and
15
N-{
1
H} NOE experiments that characterize
subnanosecond motions, CPMG and spin-lock relaxation experiments provide
quantitative information about milli- to microsecond time scale motions. In this
section, the relaxation dispersion is first defined, and, subsequently, CPMG and
spin-lock
R
2
dispersion experiments that have recently been developed for
applications to proteins are reviewed.
4.1 Relaxation Dispersion in General
Although currently the term “relaxation dispersion” or “
R
2
dispersion” often refers
to CPMG or spin-lock (off-resonance or resonance
R
1
r
) measurements, more
generally the term refers to the relaxation rates measured as a function of magnetic
field strength. Typically either the static field,
B
0
, provided by the spectrometer
magnetic or the radio-frequency (RF) field,
B
1
, generated by the probe transmitter
coil is varied over a wide range.
B
0
-dependent dispersion studies are also known as
“NMR relaxometry,” “field cycling,” or “nuclear magnetic relaxation dispersion
(NMRD)” in the literature. In these experiments
R
1
of particular nucleus is mea-
sured as a function of
B
0
[
80
-
90
]. In these relaxation dispersion applications,
the spectral density function
J
(
, allowing
various dynamical features of macromolecules, such as paramagnetic interaction
with proteins and residence times of water molecules in proteins to be obtained
[
84
,
87
]. The greatest advantage of the
R
1
dispersion experiments is that the
effective field strength varied is very wide. However, there are significant technical
challenges to varying rapidly the static field strength of the samples and to increas-
ing sensitivity [
91
-
93
]. In contrast, the dependence on
R
2
on
B
1
is readily measured,
and has been used to study chemical exchange for a long time [
94
-
101
]. The
R
2
dispersion experiment can detect such low field effects of the exchange in chemical
shifts whereas the range of variable effective field strength in the
R
2
dispersion is
relatively small compared to that of the
R
1
dispersion.
R
2
dispersion experiments
that are recently applied for biological systems will be described below.
o
) is determined at numerous values of
o
