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STD NMR 1 J* C1 h( t. I& g$ @
STD NMR - v/ I. c! R" m4 T, ^3 S1 T* W/ b
experiments detect magnetization that is transferred from a receptor protein
/ q( Z; s+ l0 `/ @5 \ to a bound ligand. Only bound ligands show STD effects. The experiment may be
9 d( j# W3 ]* d s combined with virtually any other NMR experiment, and therefore is well suitable 9 _* r2 B E9 @) {: G1 H8 m6 M
to tackle even very complex problems. In particular, in combination with multidimensional
) Q. y& p& w2 ]( D. d$ G. Q- d& o NMR a full characterization of a bound ligand out of a mixture is straightforward.
7 ?0 v7 @7 `4 }! L9 S1 b. V# B1 z9 \; d# W STD NMR is extremely robust and gives maximal effects at protein to ligand ratios
9 C/ d& T4 ?8 w$ B/ V! N/ X, G greater than ca. 1:100. It follows that less than 1 nmol of protein is necessary " P# W7 G+ M& P. h+ V s
for screening. With the availability of so called cryo probes it will be possible
; a- T. B8 v1 A3 t. P0 x# T/ f to work with hundred pmol amounts of protein. The dissociation constant should
) o: z3 h1 s! g. `' a: O; Q be in the range between nM and mM. Therefore, STD NMR covers at least two orders
; R0 Q; J! p- }7 F3 z0 b+ }1 p of magnitudes more for dissociation constants than trNOE experiments. From competitive
; N* x$ c! P. f- [* m3 B! G/ M STD experiments dissociation constants may be derived.
; i( N& ?2 `, F
8 [9 O0 o( [7 W* ]8 ]1 l) q
: f; F+ n5 [5 o0 m+ {2 Y
Schematic
' _* M4 j9 t1 {/ B" ^& B display of the STD NMR effect. Saturation of the protein leads to a direct saturation
& _2 r1 v0 a H. r; I4 n of those parts of ligand(s) in direct contact to the protein. By exchange between $ C% a: \9 Q6 l/ R1 P; y
bound and free state the saturation is transported to solution and detected & ?7 _$ }6 B! K. Q) n; y
by subtracting a spectrum with saturation from a normal spectrum.
5 X) [4 O% F7 U. o4 s STD NMR gives precise information about the binding epitope of the ligand. This
/ x% F6 M- K7 x/ b is very important information for the design of a potent drug. The optimal drug
% ~2 l) v; A9 G7 u3 X: [3 W is of optimal size and optimal shape. The size is deduced from STD NMR, and
& ^7 ?8 q9 }2 v& O: M! C6 N! X the shape is delivered by trNOE experiments.
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发表于 2010-10-18 18:42:26
