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A Fluorescence Polarization Activity-Based Protein Profiling Assay in the Discovery of Potent, Selective Inhibitors for Human Nonlysosomal Glucosylceramidase

DOI: 10.1021/jacs.7b07352 DOI Help

Authors: Daniël Lahav (Leiden University) , Bing Liu (Leiden University) , Richard J. B. H. N. Van Den Berg (Leiden University) , Adrianus M. C. H. Van Den Nieuwendijk (Leiden University) , Tom Wennekes (Leiden University) , Amar T. Ghisaidoobe (Leiden University) , Imogen Breen (The University of York) , Maria J. Ferraz (Leiden University) , Chi-lin Kuo (Leiden University) , Liang Wu (The University of York) , Paul P. Geurink (Leiden University Medical Center) , Huib Ovaa (Leiden University Medical Center) , Gijsbert A. Van Der Marel (Leiden University) , Mario Van Der Stelt (Leiden University) , Rolf G. Boot (Leiden University) , Gideon J. Davies (The University of York) , Johannes M. F. G. Aerts (Leiden University) , Herman S. Overkleeft (Leiden University)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Journal Of The American Chemical Society

State: Published (Approved)
Published: September 2017
Diamond Proposal Number(s): 13587

Open Access Open Access

Abstract: Human nonlysosomal glucosylceramidase (GBA2) is one of several enzymes that controls levels of glycolipids and whose activity is linked to several human disease states. There is a major need to design or discover selective GBA2 inhibitors both as chemical tools and as potential therapeutic agents. Here, we describe the development of a fluorescence polarization activity-based protein profiling (FluoPol-ABPP) assay for the rapid identification, from a 350+ library of iminosugars, of GBA2 inhibitors. A focused library is generated based on leads from the FluoPol-ABPP screen and assessed on GBA2 selectivity offset against the other glucosylceramide metabolizing enzymes, glucosylceramide synthase (GCS), lysosomal glucosylceramidase (GBA), and the cytosolic retaining β-glucosidase, GBA3. Our work, yielding potent and selective GBA2 inhibitors, also provides a roadmap for the development of high-throughput assays for identifying retaining glycosidase inhibitors by FluoPol-ABPP on cell extracts containing recombinant, overexpressed glycosidase as the easily accessible enzyme source.

Subject Areas: Chemistry, Biology and Bio-materials


Instruments: I03-Macromolecular Crystallography

Documents:
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