Biomedical Engineering Reference
In-Depth Information
using the starting conditions
½ P t~0 ~P tot
½ L t~0 ~L tot
½ PL t~0 ~0
ð 10 : 5 Þ
Ligand-binding events are biological switches for turning functional
processes on and off. For single-site binding, the switch is 'off' (,10%) when
[L] ,K D /10, and is 'on' (.90%) when [L] .K D 6 10. In other words, the
switch operates in three log units of concentration. Nature has found ways to
make
the
switch
more
sensitive
(positive
co-operativity)
or
less
sensitive
(negative co-operativity).
10.1.2 Ligand Binding and NMR
Many reversible biological 'transient' processes, such as signal transduction,
involve interactions with affinities weaker than 1 mM. Often macromolecules
interact using more than one binding surface. These 'polyvalent' interactions
can be quite tight, while the individual binding interfaces can be weak with mM
to mM K D values. NMR is a good tool for monitoring ligand-binding events
with affinities weaker than 1 mM. As such it is complementary to the two other
popular methods that monitor ligand binding: isothermal calorimetry (ITC)
and surface plasmon resonance (SPR, with the trade name Biacore), which
both perform best for binding that is tighter than 1 mM.
The effect of chemical exchange on the NMR signals is described by the
Bloch-McConnell equations 3,4
which for two-site exchange for a resonance
associated with the ligand read:
dM x
dt ~v L M y {R 2 M x {p L?PL M x zp PL?L M PL
x
dM y
dt ~{v L M x {R 2 M y {p L?PL M y zp PL?L M PL
y
ð 10 : 6 Þ
dM PL
x
dt
~v PL M P y {R PL
M P x {p PL?L M P x zp L?PL M x
2
dM PL
y
dt
~{v PL M P x {R PL
M P y {p PL?L M P y zp L?PL M PL
2
y
In these equations, M x and M y are the components of the coherences along
the rotating frame axes x and y, respectively, independently defined for the free
and bound ligand. v L and v PL are the resonance frequencies of the two sites,
and the R 2 values are the transverse relaxation times (linewidths). The rates
p i?j are the probabilities (in sec 21 ) that species i changes into species j.
Search WWH ::




Custom Search