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STD NMR 5 J: `+ v9 k5 v# h% G% a0 n
STD NMR
, a1 S' p4 A. \. t. |$ D experiments detect magnetization that is transferred from a receptor protein 7 n1 J7 M# x$ Y- L% `0 E9 [) Z7 M
to a bound ligand. Only bound ligands show STD effects. The experiment may be 7 k2 M- V6 p% i' F( A$ K
combined with virtually any other NMR experiment, and therefore is well suitable 6 j4 b$ {0 A- A, [
to tackle even very complex problems. In particular, in combination with multidimensional . \% Q4 f! {0 A) h
NMR a full characterization of a bound ligand out of a mixture is straightforward. 9 U( R* ?" [; i8 d
STD NMR is extremely robust and gives maximal effects at protein to ligand ratios
: W7 W6 ]& @0 h7 v4 X1 ?* M# f8 q greater than ca. 1:100. It follows that less than 1 nmol of protein is necessary * b/ t, l* V* U6 i3 h- j
for screening. With the availability of so called cryo probes it will be possible
# O9 U# p$ g! ]! Q, B, f2 | to work with hundred pmol amounts of protein. The dissociation constant should ) S- y% v' O4 l: Q4 d3 Q; |) Q
be in the range between nM and mM. Therefore, STD NMR covers at least two orders 8 h- b7 u; o# {$ P
of magnitudes more for dissociation constants than trNOE experiments. From competitive
0 c# D4 V; ]8 z. u$ g$ x+ u Z5 W STD experiments dissociation constants may be derived.
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) s- j; e! U$ |; a+ D8 |8 t
* G' g4 [+ x9 Q0 p, Q* v7 x Schematic & ^5 w* t: I2 F4 D1 |
display of the STD NMR effect. Saturation of the protein leads to a direct saturation ! ~" X+ P1 a5 B' Z: ^" U8 W
of those parts of ligand(s) in direct contact to the protein. By exchange between : ]( }; y0 Q# F8 J. Q* \0 ?
bound and free state the saturation is transported to solution and detected " _( h2 \# ^; t
by subtracting a spectrum with saturation from a normal spectrum.
' w% K8 v. ?6 ^* g" s# ?( G. a$ | STD NMR gives precise information about the binding epitope of the ligand. This . H+ \- Z* x4 U: o
is very important information for the design of a potent drug. The optimal drug
. z' r+ @8 a0 Y1 l% Z1 f) y is of optimal size and optimal shape. The size is deduced from STD NMR, and
- ~; I+ n, q2 X( J the shape is delivered by trNOE experiments.
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发表于 2010-10-18 18:42:26
