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Title: Bioluminescence-based reporters for characterizing inhibitors and activators of human Sonic Hedgehog protein autoprocessing in live cells at high throughput
Authors: Ciulla, Daniel A
Keywords: phát quang sinh học; mô tả; chất ức chế; protein; tế bào sống
Issue Date: 2023
Publisher: bioRxiv
Abstract: The Sonic hedgehog (SHh) precursor protein undergoes biosynthetic autoprocessing to cleave off and cholesterylate the SHh signaling ligand, a vital morphogen and oncogenic effector protein. Autoprocessing is self-catalyzed by SHhC, the SHh precursor’s enzymatic domain. Here we describe the development and validation of the first cellular reporter to monitor human SHhC autoprocessing non-invasively in high-throughput compatible plates. The assay couples intracellular SHhC autoprocessing to the extracellular secretion of the bioluminescent nanoluciferase enzyme. We developed a wild-type (WT) SHhC reporter line for evaluating potential autoprocessing inhibitors by concentration response-dependent suppression of extracellular bioluminescence. A conditional mutant SHhC (D46A) reporter line was developed for identifying potential autoprocessing activators by a concentration response-dependent gain of extracellular bioluminescence.
URI: http://dlib.hust.edu.vn/handle/HUST/23538
Link item primary: https://www.biorxiv.org/content/10.1101/2022.06.27.497760v1.full.pdf+html
Appears in Collections:OER - Kỹ thuật hóa học; Công nghệ sinh học - Thực phẩm; Công nghệ môi trường
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