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Assay Drug Dev Technol. 2010 Feb;8(1):37-46. doi: 10.1089/adt.2009.0219.

High-throughput multiplex flow cytometry screening for botulinum neurotoxin type a light chain protease inhibitors.

Author information

1
The Center for Molecular Discovery and Department of Pathology, University of New Mexico, Albuquerque, New Mexico 87131, USA. msaunders@salud.unm.edu

Abstract

Given their medical importance, proteases have been studied by diverse approaches and screened for small molecule protease inhibitors. Here, we present a multiplexed microsphere-based protease assay that uses high-throughput flow cytometry to screen for inhibitors of the light chain protease of botulinum neurotoxin type A (BoNTALC). Our assay uses a full-length substrate and several deletion mutants screened in parallel to identify small molecule inhibitors. The use of multiplex flow cytometry has the advantage of using full-length substrates, which contain already identified distal-binding elements for the BoNTALC, and could lead to a new class of BoNTALC inhibitors. In this study, we have screened 880 off patent drugs and bioavailable compounds to identify ebselen as an in vitro inhibitor of BoNTALC. This discovery demonstrates the validity of our microsphere-based approach and illustrates its potential for high-throughput screening for inhibitors of proteases in general.

PMID:
20035615
PMCID:
PMC3096553
DOI:
10.1089/adt.2009.0219
[Indexed for MEDLINE]
Free PMC Article
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