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J Am Chem Soc. 2018 May 9;140(18):5914-5924. doi: 10.1021/jacs.7b13003. Epub 2018 Apr 30.

β-Glucocerebrosidase Modulators Promote Dimerization of β-Glucocerebrosidase and Reveal an Allosteric Binding Site.

Author information

1
Department of Neurology , Northwestern University Feinberg School of Medicine , Chicago , Illinois 60611 , United States.
2
Department of Chemistry, Department of Molecular Biosciences, Chemistry of Life Processes Institute, Center for Molecular Innovation and Drug Discovery, and Center for Developmental Therapeutics , Northwestern University , Evanston , Illinois 60208 , United States.
3
Department of Chemistry and Molecular Biosciences, and Proteomics Center of Excellence , Northwestern University , Evanston , Illinois 60208 , United States.
4
Department of Molecular Biosciences , Northwestern University , Evanston , Illinois 60208 , United States.
5
Lysosomal Therapeutics Inc. , 19 Blackstone Street , Cambridge , Massachusetts 02139 , United States.
6
Proteros Biostructures GmbH , Bunsenstrasse 7a , Martinsried 82152 , Germany.
7
Department of Pediatrics , Northwestern University Feinberg School of Medicine , Chicago , Illinois 60611 , United States.

Abstract

β-Glucocerebrosidase (GCase) mutations cause Gaucher's disease and are a high risk factor in Parkinson's disease. The implementation of a small molecule modulator is a strategy to restore proper folding and lysosome delivery of degradation-prone mutant GCase. Here, we present a potent quinazoline modulator, JZ-4109, which stabilizes wild-type and N370S mutant GCase and increases GCase abundance in patient-derived fibroblast cells. We then developed a covalent modification strategy using a lysine targeted inactivator (JZ-5029) for in vitro mechanistic studies. By using native top-down mass spectrometry, we located two potentially covalently modified lysines. We obtained the first crystal structure, at 2.2 Å resolution, of a GCase with a noniminosugar modulator covalently bound, and were able to identify the exact lysine residue modified (Lys346) and reveal an allosteric binding site. GCase dimerization was induced by our modulator binding, which was observed by native mass spectrometry, its crystal structure, and size exclusion chromatography with a multiangle light scattering detector. Finally, the dimer form was confirmed by negative staining transmission electron microscopy studies. Our newly discovered allosteric site and observed GCase dimerization provide a new mechanistic insight into GCase and its noniminosugar modulators and facilitate the rational design of novel GCase modulators for Gaucher's disease and Parkinson's disease.

PMID:
29676907
PMCID:
PMC6098685
DOI:
10.1021/jacs.7b13003
[Indexed for MEDLINE]
Free PMC Article

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