Synthetic substrates specific to activated plasmin can monitor the enzymatic functional status in situ in breast cancer cells
Synthetic substrates specific to activated plasmin can monitor the enzymatic functional status in situ in breast cancer cells
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活化纤溶酶特异性的合成底物可以原位监测乳腺癌细胞中的酶功能状态
DOI：
10.1111/cbdd.12232
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发表时间：
2014
期刊：
影响因子：
--
通讯作者：
Y.
中科院分区：
文献类型：
--
作者：
Gohda;K.;Fujimori;K.;Teno;N.;Wanaka;K.;and Tsuda;Y.
我们在此努力克服PCR和IHC等表达分析的局限性，基于靶分子和探针分子之间的分子识别，通过设计针对靶分子的合成底物来直接原位估计酶功能。特异性底物包含探测单元（用于特异性酶结合的有机片段）和反应单元（受催化作用的天然肽）。在这项研究中，使用由其抑制剂设计的纤溶酶特异性合成底物在 MDA-MB231 乳腺癌细胞中检查了纤溶酶原向纤溶酶的激活。通过荧光团与特定底物同时结合，可以成功地观察到激活的纤溶酶的定位和功能。这将是首次通过现场直接观察激活纤溶酶。我们直接监测目标酶功能的概念可以直接用于其他蛋白酶，例如组织蛋白酶或半胱天冬酶。此外，这种底物概念作为“定制底物”将被用作体内新型功能性分子探针，并具有适当的可检测探针。
We here strove to overcome the limitations of expression analyses such as PCR and IHC, based on molecular recognition between target and probe molecules, by designing synthetic substrates specific to the target molecules to directly estimate the enzymatic functionalityin situ. The specific substrate contains a probing unit, which is an organic fragment for specific enzyme binding, and a reactive unit, which is a natural peptide subject to catalysis. In this study, the activation of plasminogen to plasmin was examined in MDA‐MB231 breast cancer cells using the plasmin‐specific synthetic substrates designed from their inhibitors. The localization and function of the activated plasmin were successfully visualized by fluorophore combined with the specific substrate concurrently. This would be the first time for activated plasmin at workin situby direct observation. Our concept to directly monitor the functionality of target enzymes can be used straightforwardly for other proteases such as cathepsins or caspases. Also, this substrate concept as a ‘tailor‐made substrate’ would be utilized as a novel functional molecular probein vivowith appropriate detectable probes.