A cellular
system for positive selection of beta-secretase inhibitors.
Oliver Middendorp,
Urs Luethi, Alcide Barberis
ESBATech AG, Protease Unit, Wagistr. 21, Zürich-Schlieren, 8952, Switzerland
(middendorp@esbatech.com)
We describe a
cellular selection system in yeast to identify inhibitors of the beta-secretase
BACE. The system is designed such that growth of yeast cells is stimulated upon
inhibition of BACE. Cleavage of a membrane-anchored invertase-APP fusion
protein results in liberation and secretion of the invertase moiety. Secreted
invertase catalyses the hydrolysis of sucrose to generate fructose and glucose.
The newly generated glucose dominantly represses transcription of a selectable
reporter gene under the control of the GAL1-promoter. Inhibition of BACE
impairs the release of the invertase moiety and, consequently, no glucose is
generated. In the absence of glucose, transcription of the cell growth marker
takes place and the cells can grow under selective conditions.