弹性蛋白酶抑制剂筛选用蛋白质芯片和化合物阵列的设计与应用

Design and application of protein chip and chemical arrays used for screening elastase inhibitors

介绍了用于筛选弹性蛋白酶抑制剂的化合物阵列（药物筛选生物芯片）的设计与应用。这种化合物阵列芯片由活性蛋白酶膜、化合物阵列及底物微阵构成。其主要原理是弹性蛋白酶与显色底物反应之后，可以催化底物转化为显色产物，根据颜色的差异，可以确定化合物是否对酶活性具有抑制作用以及抑制作用的强弱。为提高检测灵敏度，在反应系统中加入了与反应产物形成颜色反差的生物指示剂溴酚蓝作为背景。当酶与底物反应时，底物转化为黄色的产物，在背景色中呈现绿色；当酶活性被化合物抑制时，无黄色底物生成，点样点呈背景蓝色。使用该化合物微阵列方法对11680个化合物进行了筛选，发现了两个抑制作用较强的化合物，并通过常规的微孔板方法证明了筛选结果的准确性。说明化合物阵列蛋白质芯片方法是有效、灵敏的高通量筛选方法。

A chemical array has been established to screen enzymatic inhibitors. The chip includes an active proteinase film, screened chemical arrays distributed on the film and substrate microarrays. The principle of detecting is that the elastase hydrolysises the synthesized chromogenic substrates and turns them into chromogenic products which can test if the chemical reduces the enzymatic activity by detecting the color change of the spots. In order to facilitate the detection process, the bromehlorphenol blue（BPB） was added into the substrate solutions. When elastase retained its activity, yellow was produced by products combined with blue of BPB and the spots showed green. When enzymatic activity was reduced, substrates could not be entirely turned into yellow products and blue was enhanced. At last, 11680 compounds were screened and two active compounds were hit, which illuminated that this assay is effective, sensitive and applicable to high throughput screening.