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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 C% g6 b) g3 r( E experiments detect magnetization that is transferred from a receptor protein , @5 d g1 b5 P' R/ J1 i to a bound ligand. Only bound ligands show STD effects. The experiment may be 4 {; ?4 E' o$ _# q6 Y+ b9 N0 a. j5 W# ~ combined with virtually any other NMR experiment, and therefore is well suitable 4 F8 l2 A9 o/ |& c! z7 P to tackle even very complex problems. In particular, in combination with multidimensional & g/ t4 O$ q$ q8 c$ ^+ Y NMR a full characterization of a bound ligand out of a mixture is straightforward. ! m6 q) N m4 ^- k% {; W1 Z STD NMR is extremely robust and gives maximal effects at protein to ligand ratios % `8 L z/ D. E4 h' U greater than ca. 1:100. It follows that less than 1 nmol of protein is necessary & H1 P% B7 [- n( ]2 u for screening. With the availability of so called cryo probes it will be possible 5 Y/ B$ J( H1 L# k; x! V3 B8 u6 R to work with hundred pmol amounts of protein. The dissociation constant should % y7 G) c6 P( \8 a: @0 W9 a be in the range between nM and mM. Therefore, STD NMR covers at least two orders ; a4 b# v9 l8 [ of magnitudes more for dissociation constants than trNOE experiments. From competitive - e7 |& _6 z0 x STD experiments dissociation constants may be derived.

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' A2 v+ \% w! a/ {: J/ B3 P6 x8 u Schematic # m; Y/ f9 w! y5 `- J display of the STD NMR effect. Saturation of the protein leads to a direct saturation # |3 Q4 P& |5 {) ^ of those parts of ligand(s) in direct contact to the protein. By exchange between . r- d/ M7 t* c$ s/ q' o2 u* o bound and free state the saturation is transported to solution and detected 7 ]5 J' O4 V8 n! Y& D8 b5 ` v9 Y+ m4 } by subtracting a spectrum with saturation from a normal spectrum.
, s# K0 |# I( f8 a STD NMR gives precise information about the binding epitope of the ligand. This + |& h' U) [" a is very important information for the design of a potent drug. The optimal drug 5 j7 m1 }4 c9 A. h$ e( H: [ is of optimal size and optimal shape. The size is deduced from STD NMR, and 0 f/ V, j6 z6 P: e the shape is delivered by trNOE experiments.

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