晶片上蛋白質間作用力之探討

Study of Protein-proteinInteraction on Chip

陳惠民 ； 高豐生

《國家奈米元件實驗室奈米通訊》 18卷4期 (2011/12) Pp. 20-24

https://doi.org/10.29913/NDLC.201112.0004

摘要

在我們過去的研究中（Dong, G－C., et al., J. Med. Chem. 2006, 49, 1845-1854; Dong, G－C., et al.,Pharmaceutical Res. 2009, 26, 375-381; C－C. Chen, et al., 2011, Fitoterapia, 82, 1249－1257） 指出利用表面電漿共振（Surface Plasma Resonance; SPR）為檢測之工具，用結合蛋白質固定在晶片的方法，可以用來篩選標靶藥物。然而，藉由量測標靶藥物結合在蛋白質上的折射角度不同所換算出光量之差異，來判斷藥物附著在蛋白質上之有效性，是比較間接及不精確的方法。在這篇文章中，我們利用蛋白質間作用力差異之量測，作為更直接及準確分析方式之藥物篩選。成功的範例是，我們將T細胞表面的接受器（CD28）與B細胞表面的接受器（CD80），分別固定在晶片表面（imm-CD28）與原子力顯微鏡（AFM）探針的表面上（afm-CD80），並且加入免疫抑制劑cynarin，來探討其對於兩者間作用力的影響。 結果顯示出，若是加入Cynarin，則imm-CD28 與afm-CD80間的結合機率（Binding Probability），從原本的61 ± 5% 降至47 ± 6 %。 而平均的分離力（unbinding force）則從61.4 pN降至38.9 pN，減少了約37 %。此結果與以往以SPR的量測結果（僅約9 %之差異），在定量差異上有明顯的提升。因此，以AFM作為藥物篩選的檢測工具或蛋白質間作用力之量測，可以大大提升其準確性及應用性。

關鍵字

原子力顯微鏡 ； 蛋白質間作用力 ； 分離力 ； 藥物篩選

並列摘要

Drug screening based on the binding of drug target to the immobilization of protein (disease related receptor) on chip via surface plasma resonance (SPR) as detection probe has been reported by us previously (Dong, G-C., et al., J. Med. Chem. 2006, 49, 1845-1854; Dong, G-C., et al., Pharmaceutical Res. 2009, 26, 375-381; C-C. Chen, et al., 2011, Fitoterapia, 82, 1249-1257). However, the measurement of binding strength by the blocking of target drug to protein-protein interaction may be indirect. In this communication, we demonstrated that the direct and accurate measurement of protein-protein interaction can be done by atomic force microscopy (AFM) via immobilization of one protein on chip (imm-protein) and another on tip of AFM (afm-protein). Preliminarily, we observed the binding between T-cell receptor, CD28 (imm-CD28), and B-cell receptor, CD80 (afm-CD80), with an immuno-suppressive blocker, cynarin. The results showed that the binding events was largely reduced from 61 ± 5 % to 47 ± 6 % if cynarin was bound with CD28 .rst. Furthermore, the average unbinding forces were reduced from 61.4 pN to 38.9 pN with the blocking e.ect about 37 % as compared with about 9 % of surface plasma resonance (SPR) measurement. Using AFM as a detection tool, the signi.cant quantity di.erences above caused by drug target binding therefore can be applied to an efficient and accurate drug screening or protein-protein interaction measurement.

並列關鍵字

Atomic Force Microscopy (AFM) ； Protein-proteinInteraction ； Unbinding Force ； Drug Screening