20.500.12556/RUL-164279 A novel simple fluorometric protease assay for monitoring hydrolysis of proteins in real time Measuring the activity of proteases is essential for investigating both the physiological functions and commercial applications of these enzymes. In contrast to the numerous protease assays that are based on chromogenic or fluorogenic peptide substrates, there is a lack of approaches to monitor degradation of proteins in real time. Here we report a protease assay where SYPRO Orange is employed as a fluorogenic probe to follow proteolysis. The functionality of the assay was demonstrated with the two subtilases of varying thermostability, using four different protein substrates. The assay is compatible with a real-time PCR instrument which allows continuous fluorescence measurements in low-volume samples even at high temperatures. This makes the assay especially suitable for high-throughput characterization of thermostable proteases. protease assay proteolysis subtilisin SYPRO orange fluorescence real-time PCR thermostability encimi proteaze aktivnost proteaz proteoliza PCR true false true Angleški jezik Slovenski jezik Članek v reviji 2024-10-18 12:32:07 2024-10-18 12:32:11 2024-10-19 04:25:47 0000-00-00 00:00:00 2025 0 0 9 str. art. 115688 Vol. 696 Jan. 2025 0000-00-00 Zaloznikova Objavljeno NiDoloceno 0000-00-00 0000-00-00 0000-00-00 577.152.34 0003-2697 10.1016/j.ab.2024.115688 212076803 1-s2.0-S000326972400232X-main.pdf 1-s2.0-S000326972400232X-main.pdf 1 9FD28B910E494582314C2A6E6F7898C3 d6ff5740555e9c2e079e846c2d8b24c64963ebb1673b11ed279e8d6fce29e32e 3c8da4a8-8d3c-11ef-b232-0050569b8976 https://repozitorij.uni-lj.si/Dokument.php?lang=slv&id=194575 https://www.sciencedirect.com/science/article/pii/S000326972400232X 0 fa614c66-8d3b-11ef-b232-0050569b8976 https://repozitorij.uni-lj.si/Dokument.php?lang=slv&id=194574 Biotehniška fakulteta 0 0 0