SPR設備在大分子復合物活化位點結合反應分析中的應用
ACS Chem. Biol. 2013, 8, 2794−2801 Michael P. Storz, ect.
美國化學生物學，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
對PqsD抑制劑手性異構體的生物化學和生物物理學分析揭示了其穩(wěn)定結合反應和與其具有明顯相反熱動力學特性的旋光異構體的存在
Binding Site Analysis by SPR. To investigate whether our compounds bind in the active site, we performed binding experiments using SPR.
SPR分析結合位點 為了探究被檢復合物是否在活化位點結合，進行了利用SPR技術的結合實驗。
Surface Plasmon Resonance (SPR) Spectrometry. SPR binding studies were performed using a Reichert SR7500DC instrument optical biosensor (Reichert Technologies)。
表面等離子共振光譜儀 SPR結合實驗使用了Reichert SR7500DC 光學傳感儀（Reichert Technologies，新型號為 2SPR）
圖一
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.
圖1 利用SPR技術闡明結合位點。給天然PqsD加上抑制因子分別引起了A圖和B圖的信號曲線，分別對應復合物2和3。相對于復合物2對應的曲線（如圖C）,當抑制因子加到被ACoA預處理過的PqsD時沒有出現(xiàn)復合物3應有的反應信號曲線（如圖D）。
Figure S9: Binding site analysis of (R)-3 and (S)-3 by SPR
圖S9：通過SPR技術對(R)-3和 (S)-3結合位點分析。
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.
圖S9，SPR實驗中分別使用了250和125μM的(R)-3 (藍) or (S)-3 (橘)。復合物加入到自然PqsD中，并且記錄下了對應的信號曲線(圖A) 和 (圖B)，當同樣濃度(R)-3 (如圖C) 和 (S)-3 (如圖D)加到被ACoA預處理過的PqsD時沒有出現(xiàn)反應信號（如圖D）。另外，當持續(xù)加入(R)-3直到SPR信號飽和，隨后再加入(S)-3也不能影響反應了（如圖E）。結果表明兩種異構體的結合位點是被鄰氨基苯甲酸鹽封閉了。
