SPR设备在大分子复合物活化位点结合反应分析中的应用
ACS Chem. Biol. 2013, 8, 2794−2801 Michael P. Storz, ect.
美国化学生物学，2013.8.2794-2801 Michael P. Storz等人
Biochemical and Biophysical Analysis of a Chiral PqsD Inhibitor Revealing Tight-binding Behavior and Enantiomers with Contrary Thermodynamic Signatures
对PqsD抑制剂手性异构体的生物化学和生物物理学分析揭示了其稳定结合反应和与其具有明显相反热动力学特性的旋光异构体的存在
Binding Site Analysis by SPR. To investigate whether our compounds bind in the active site, we performed binding experiments using SPR.
SPR分析结合位点 为了探究被检复合物是否在活化位点结合，进行了利用SPR技术的结合实验。
Surface Plasmon Resonance (SPR) Spectrometry. SPR binding studies were performed using a Reichert SR7500DC instrument optical biosensor (Reichert Technologies)。
表面等离子共振光谱仪 SPR结合实验使用了Reichert SR7500DC 光学传感仪（Reichert Technologies，新型号为 2SPR）
图一
Figure 1. Elucidation of the binding site by SPR. Addition of inhibitor to native PqsD leads to response curves (A) and (B) for compounds 2 and 3, respectively. In contrast to compound 2 (C), no response curve for 3 (D) is observed after addition to PqsD pretreated with ACoA.
图1 利用SPR技术阐明结合位点。给天然PqsD加上抑制因子分别引起了A图和B图的信号曲线，分别对应复合物2和3。相对于复合物2对应的曲线（如图C）,当抑制因子加到被ACoA预处理过的PqsD时没有出现复合物3应有的反应信号曲线（如图D）。
Figure S9: Binding site analysis of (R)-3 and (S)-3 by SPR
图S9：通过SPR技术对(R)-3和 (S)-3结合位点分析。
Figure S9. SPR experiment using 250 and 125μM of (R)-3 (blue) or (S)-3 (orange), respectively. Compounds were added to native PqsD and response curves (A) and (B) were recorded. No response was observed when identical concentrations of (R)-3 (C) or (S)-3 (D) were added to PqsD pretreated with ACoA. Furthermore, when (R)-3 was added until saturation of the SPR signal, subsequent addition of (S)-3 did not affect the observed response (E). The results indicate that the binding sites of both enantiomers are blocked by anthranilate.
图S9，SPR实验中分别使用了250和125μM的(R)-3 (蓝) or (S)-3 (橘)。复合物加入到自然PqsD中，并且记录下了对应的信号曲线(图A) 和 (图B)，当同样浓度(R)-3 (如图C) 和 (S)-3 (如图D)加到被ACoA预处理过的PqsD时没有出现反应信号（如图D）。另外，当持续加入(R)-3直到SPR信号饱和，随后再加入(S)-3也不能影响反应了（如图E）。结果表明两种异构体的结合位点是被邻氨基苯甲酸盐封闭了。
