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STD NMR $ g) j0 m' y( x% n8 z( M9 j
STD NMR 7 H, o" g+ A- k9 u* Q: l
experiments detect magnetization that is transferred from a receptor protein & T! q8 Y" \8 V9 W5 K/ A
to a bound ligand. Only bound ligands show STD effects. The experiment may be
9 K, [" T$ Z0 ^0 b combined with virtually any other NMR experiment, and therefore is well suitable
& q9 ^4 t; J3 ~. f to tackle even very complex problems. In particular, in combination with multidimensional - D0 U* T( g; H9 U. o3 p
NMR a full characterization of a bound ligand out of a mixture is straightforward. 8 e& X! S' A4 y1 l: N5 c
STD NMR is extremely robust and gives maximal effects at protein to ligand ratios
/ C7 X0 ~8 L; Z1 N; W! n greater than ca. 1:100. It follows that less than 1 nmol of protein is necessary
; ? p2 }8 g: p) y( ` for screening. With the availability of so called cryo probes it will be possible * ?- F ]4 x2 r, `1 h8 |
to work with hundred pmol amounts of protein. The dissociation constant should
& r+ y& S5 w: L; h be in the range between nM and mM. Therefore, STD NMR covers at least two orders & i. F' F* Q- G; \' ?, k1 n
of magnitudes more for dissociation constants than trNOE experiments. From competitive 1 ^4 K( [1 z7 c" c+ b
STD experiments dissociation constants may be derived.
% o1 N/ C5 K* c/ D
6 p& ]. w; j& l# d6 D! f
% c* A0 W0 j6 |' z4 r" t$ w& }: e
Schematic
: [6 n0 K2 J+ q( V display of the STD NMR effect. Saturation of the protein leads to a direct saturation ( D# d3 H7 B2 S% i( [
of those parts of ligand(s) in direct contact to the protein. By exchange between ) }1 G; {, O$ k* P8 e. U" q1 p
bound and free state the saturation is transported to solution and detected % D4 `& X* V* r$ q) f# @
by subtracting a spectrum with saturation from a normal spectrum.
1 ?7 W7 t" }/ ]1 ~6 e" Y STD NMR gives precise information about the binding epitope of the ligand. This # n* z5 C5 Z$ Z
is very important information for the design of a potent drug. The optimal drug
* s9 i$ [) g( y' h9 k is of optimal size and optimal shape. The size is deduced from STD NMR, and / |, j4 n5 f% L; w
the shape is delivered by trNOE experiments.
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发表于 2010-10-18 18:42:26
