Selective labelling of GBA2 in cells with fluorescent β-d-arabinofuranosyl cyclitol aziridines

TY - JOUR

T1 - Selective labelling of GBA2 in cells with fluorescent β-d-arabinofuranosyl cyclitol aziridines

AU - Su, Qin

AU - Louwerse, Max

AU - Lammers, Rob F.

AU - Maurits, Elmer

AU - Janssen, Max

AU - Boot, Rolf G.

AU - Borlandelli, Valentina

AU - Offen, Wendy A.

AU - Linzel, Daniël

AU - Schröder, Sybrin P.

AU - Davies, Gideon J.

AU - Overkleeft, Herman S.

AU - Artola, Marta

AU - Aerts, Johannes M.F.G.

N1 - Publisher Copyright: © 2024 The Royal Society of Chemistry.

PY - 2024/9/3

Y1 - 2024/9/3

N2 - GBA2, the non-lysosomal β-glucosylceramidase, is an enzyme involved in glucosylceramide metabolism. Pharmacological inhibition of GBA2 by N-alkyl iminosugars is well tolerated and benefits patients suffering from Sandhoff and Niemann-Pick type C diseases, and GBA2 inhibitors have been proposed as candidate-clinical drugs for the treatment of parkinsonism. With the ultimate goal to unravel the role of GBA2 in (patho)physiology, we sought to develop a GBA2-specific activity-based probe (ABP). A library of probes was tested for activity against GBA2 and the two other cellular retaining β-glucosidases, lysosomal GBA1 and cytosolic GBA3. We show that β-d-arabinofuranosyl cyclitol aziridine (β-d-Araf aziridine) reacts with the GBA2 active site nucleophile to form a covalent and irreversible bond. Fluorescent β-d-Araf aziridine probes potently and selectively label GBA2 both in vitro and in cellulo, allowing for visualization of the localization of overexpressed GBA2 using fluorescence microscopy. Co-staining with an antibody selective for the lysosomal β-glucosylceramidase GBA1, shows distinct subcellular localization of the two enzymes. We proffer our ABP technology for further delineating the role and functioning of GBA2 in disease and propose the β-d-Araf aziridine scaffold as a good starting point for the development of GBA2-specific inhibitors for clinical development.

AB - GBA2, the non-lysosomal β-glucosylceramidase, is an enzyme involved in glucosylceramide metabolism. Pharmacological inhibition of GBA2 by N-alkyl iminosugars is well tolerated and benefits patients suffering from Sandhoff and Niemann-Pick type C diseases, and GBA2 inhibitors have been proposed as candidate-clinical drugs for the treatment of parkinsonism. With the ultimate goal to unravel the role of GBA2 in (patho)physiology, we sought to develop a GBA2-specific activity-based probe (ABP). A library of probes was tested for activity against GBA2 and the two other cellular retaining β-glucosidases, lysosomal GBA1 and cytosolic GBA3. We show that β-d-arabinofuranosyl cyclitol aziridine (β-d-Araf aziridine) reacts with the GBA2 active site nucleophile to form a covalent and irreversible bond. Fluorescent β-d-Araf aziridine probes potently and selectively label GBA2 both in vitro and in cellulo, allowing for visualization of the localization of overexpressed GBA2 using fluorescence microscopy. Co-staining with an antibody selective for the lysosomal β-glucosylceramidase GBA1, shows distinct subcellular localization of the two enzymes. We proffer our ABP technology for further delineating the role and functioning of GBA2 in disease and propose the β-d-Araf aziridine scaffold as a good starting point for the development of GBA2-specific inhibitors for clinical development.

UR - http://www.scopus.com/inward/record.url?scp=85203202417&partnerID=8YFLogxK

U2 - 10.1039/d3sc06146a

DO - 10.1039/d3sc06146a

M3 - Article

AN - SCOPUS:85203202417

SN - 2041-6520

VL - 15

SP - 15212

EP - 15220

JO - Chemical Science

JF - Chemical Science

IS - 37

ER -