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[求助]NMR检测小分子配体与蛋白相互作用

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发表于 2010-10-18 18:42:26 | 显示全部楼层 |阅读模式

刚接触NMR,想用来检测一下一种三个苯环的小分子配体与一种三万KDa的蛋白的相互作用,

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请教各位大侠,是做小分子的H谱呢?还是做大分子的H谱?

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做的话对于溶剂有什么要求吗?蛋白用什么溶剂呢?小分子用DMSO溶解可以吗?

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各位大侠帮帮忙,不胜感激啊

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发表于 2010-10-18 23:59:29 | 显示全部楼层
应该是用std 实验吧。
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 楼主| 发表于 2010-10-19 12:22:57 | 显示全部楼层
什么事STD实验啊?大侠能详细解释一下吗?小弟初次接触NMR
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发表于 2010-10-19 20:45:31 | 显示全部楼层
同问。蛋白的核磁咋做?
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发表于 2010-10-19 23:54:37 | 显示全部楼层
 

STD NMR

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STD NMR 1 f+ I0 P U+ a% r- z2 ?& ` experiments detect magnetization that is transferred from a receptor protein " T' _6 B# W- X4 a! J o: U2 C+ _ to a bound ligand. Only bound ligands show STD effects. The experiment may be 7 t4 |9 i: M8 C5 K! I% | combined with virtually any other NMR experiment, and therefore is well suitable ) f+ W) v( ~) p4 l2 Y& v to tackle even very complex problems. In particular, in combination with multidimensional * v5 R, j" r& [! x( P0 M4 S( ]0 f NMR a full characterization of a bound ligand out of a mixture is straightforward. 0 C1 L. u% Q8 @ STD NMR is extremely robust and gives maximal effects at protein to ligand ratios ( J6 p$ X9 o" |9 S3 U greater than ca. 1:100. It follows that less than 1 nmol of protein is necessary 9 b* t, ]% [7 w. J3 U4 E' @0 s for screening. With the availability of so called cryo probes it will be possible ) s3 l" J, d5 W# H+ k% o s to work with hundred pmol amounts of protein. The dissociation constant should / m1 B9 v5 w, v5 ~3 w$ K. E+ R be in the range between nM and mM. Therefore, STD NMR covers at least two orders / y* U0 M( ]1 {) x4 K of magnitudes more for dissociation constants than trNOE experiments. From competitive & @6 u, {* _) X; k3 x STD experiments dissociation constants may be derived.

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_& G3 a2 d7 Z/ v9 z- I [ Schematic 8 n. x! z5 r' r: L display of the STD NMR effect. Saturation of the protein leads to a direct saturation 4 x* U" } K, S. O5 C4 P& S% i( t% Q0 q of those parts of ligand(s) in direct contact to the protein. By exchange between " x* t9 T9 K" R; c" O) s bound and free state the saturation is transported to solution and detected ' R+ M. _) ^) C* ]2 ~ ~( R F2 z by subtracting a spectrum with saturation from a normal spectrum.
; @) t! t, b4 b) s- B STD NMR gives precise information about the binding epitope of the ligand. This 0 f0 f9 | M6 Q( |: V is very important information for the design of a potent drug. The optimal drug - }. f' S" N# v! e is of optimal size and optimal shape. The size is deduced from STD NMR, and " k3 I N9 [# J the shape is delivered by trNOE experiments.

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