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Yeast-hybrid based high-throughput assay for identification of anthrax lethal factor inhibitors

Title
Yeast-hybrid based high-throughput assay for identification of anthrax lethal factor inhibitors
Author
윤문영
Keywords
Anthrax lethal factor; Cell-based high-throughput assay; Chemical library screening; Yeast
Issue Date
2011-01
Publisher
Elsevier
Citation
Biochemical and Biophysical Research Communications, 2011, 404(1), P.517-522
Abstract
Inhibitors of anthrax lethal factor (LF) are currently being sought as effective therapeutics for the treatment of anthrax. Here we report a novel screening approach for inhibitors of LF, a yeast-hybrid-based assay system in which the expression of reporter genes from a Gal4 promoter is repressed by LF proteolytic activity. Yeast cells were co-transformed with LF and a chimeric transcription factor that contains an LF substrate sequence inserted between the DNA-binding and activation domains of Gal4. In the resulting yeast cells, LF cleaves the substrate, thus inactivating the chimeric Gal4 and resulting in lack of expression of reporter genes. Compounds that inhibit LF cleavage of its substrate are identified by changes in reporter gene activity. Relative to in vitro screens for inhibitors of LF proteolytic activity, this screen has the advantage of excluding compounds that are toxic or non-permeable to eukaryotic cells. Additionally, the screen has the advantage of being fast, easy and cheap because exogenous LF and substrate are not needed. An initial chemical library screen with this system has identified four candidate inhibitors which were confirmed to inhibit LF protease activity in an in vitro assay. Furthermore, FBS-00831, one of the compounds identified, protects Raw 264.7 macrophages from anthrax lethal toxin and the possible binding site on LF was also evaluated by molecular docking.
URI
https://www.sciencedirect.com/science/article/pii/S0006291X10022400?via%3Dihub
ISSN
0006-291X; 1090-2104
DOI
10.1016/j.bbrc.2010.12.015
Appears in Collections:
COLLEGE OF NATURAL SCIENCES[S](자연과학대학) > CHEMISTRY(화학과) > Articles
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