A novel cell-based sensor detecting the activity of individual basic proprotein convertases [article]

Karin Löw, Kornelia Hardes, Chiara Fedeli, Nabil G. Seidah, Daniel B. Constam, Antonella Pasquato, Torsten Steinmetzer, Alexandre Roulin, Stefan Kunz
2018 bioRxiv   pre-print
ABSTRACTThe basic proprotein convertases (PCs) furin, PC1/3, PC2, PC5/6, PACE4, PC4, and PC7 are promising drug targets for human diseases. However, developing selective inhibitors remains challenging due to overlapping substrate recognition motifs and limited structural information. Classical drug screening approaches for basic PC inhibitors involve homogeneous biochemical assays using soluble recombinant enzymes combined with fluorogenic substrate peptides and do not accurately recapitulate
more » ... e complex cellular context of the basic PC-substrate interaction. We report here PCific, a novel cell-based molecular sensor that allows rapid screening of candidate inhibitors and their selectivity toward individual basic PCs within mammalian cells. PCific consists of Gaussia luciferase linked to a sortilin-1 membrane anchor via a cleavage motif that allows efficient release of luciferase specifically if individual basic PCs are provided in cis. Screening of selected candidate peptidomimetic inhibitors revealed that PCific can readily distinguish between general and selective PC inhibitors in a high-throughput screening format.
doi:10.1101/464032 fatcat:qjngca4np5as7l3yn6rn64n5ne