Cao Yi, Han Feng-mei, Chen Yong
Hubei Provincal Key Lab of Bio-Technology of Traditional Chinese Medicine, Hubei University, Wuhan 430062, China.
Yao Xue Xue Bao. 2007 Apr;42(4):408-12.
A new MS-titration method for the non-covalent binding of protein-ligand based on the research of berberine and alpha1-acid glycoprotein was established. The major presumption of new method is that the total concentration of protein-ligand complex is approximately the same as the total concentration of acting protein if a certain extent of affinity is existed between protein and ligand, in addition, the mole amount of acting ligand is more than that of acting protein. The non-covalent binding behaviours between berberine and alpha1-acid glycoprotein was studied by using electrospray ionization ion trap mass spectrometry (ESI-ITMS) , and the results were verified by fluorescence quenching method. The results showed that the binding behaviours between berberine and alpha1-acid glycoprotein, for example, stability constant, number of binding site, type of the main binding force, were almost the same by using the new MS-titration method and fluorescence quenching method. Comparing with the reported MS-titration method, the presented MS-titration method in this paper is more simple and applicable, does not demand much for the devices, and can lead to reliable results in same cases.
基于黄连素与α1-酸性糖蛋白的研究，建立了一种用于蛋白质-配体非共价结合的新型质谱滴定法。新方法的主要假设是，如果蛋白质与配体之间存在一定程度的亲和力，蛋白质-配体复合物的总浓度与作用蛋白的总浓度大致相同，此外，作用配体的摩尔量大于作用蛋白的摩尔量。采用电喷雾电离离子阱质谱（ESI-ITMS）研究了黄连素与α1-酸性糖蛋白之间的非共价结合行为，并通过荧光猝灭法对结果进行了验证。结果表明，黄连素与α1-酸性糖蛋白之间的结合行为，如稳定常数、结合位点数、主要结合力类型，采用新型质谱滴定法和荧光猝灭法几乎相同。与报道的质谱滴定法相比，本文提出的质谱滴定法更简单、适用，对设备要求不高，在相同情况下能得出可靠的结果。
