AJOU Central Library Repository: Development of a novel protease assay method by using an enzymatic cascades of engineered zymogens

Development of a novel protease assay method by using an enzymatic cascades of engineered zymogens

Alternative Title: Development of a novel protease assay method by using an enzymatic cascades of engineered zymogens

Alternative Abstract: Members of the matrix metalloproteinase (MMP) family have been implicated in disease states such as arthritis, periodontal disease, and tumor cell invasion and metastasis. MMPs were identified as therapeutic targets for cancers, and reviews have been published for the roles of MMPs in cancers. We previously reported a protease sensor platform developed by using an engineered ß-lactamase zymogen, and it was used for assaying MMP-2 activity. The assay method has a signal amplification step by ß-lactamase and thus showed a high sensitivity. In order to improve the method, we, in this study, have engineered a caspase-3 zymogen (pro-caspase-3) that can be activated by MMP-2 and then developed a new MMP-2 assay method by using an enzymatic cascade composed of the pro-caspase-3 enzyme and a ß-lactamase zymogen which can be activated by caspase-3. This method includes two signal amplification steps by caspase-3 and ß-lactamase and showed a detection of limit (LOD) of 29 pM, which is, to our knowledge, the lowest LOD reported so far. Two engineered zymogens can be easily engineered for other protease, and thus we believe that the signal cascade platform will be developed for assaying various proteases.

Appears in Collections:: Graduate School of Ajou University > Department of Molecular Science and Technology > 3. Theses(Master)

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