Visualization of Active Glucocerebrosidase in Rodent Brain with High Spatial Resolution following In Situ Labeling with Fluorescent Activity Based Probes

Daniela Herrera Moro Chao; Wouter W Kallemeijn; Andre R A Marques; Marie Orre; Roelof Ottenhoff; Cindy van Roomen; Ewout Foppen; Maria C Renner; Martina Moeton; Marco van Eijk; Rolf G Boot; Willem Kamphuis; Elly M Hol; Jan Aten; Hermen S Overkleeft; Andries Kalsbeek; Johannes M F G Aerts

doi:10.1371/journal.pone.0138107

Visualization of Active Glucocerebrosidase in Rodent Brain with High Spatial Resolution following In Situ Labeling with Fluorescent Activity Based Probes

Daniela Herrera Moro Chao, Wouter W Kallemeijn, Andre R A Marques, Marie Orre, Roelof Ottenhoff, Cindy van Roomen, Ewout Foppen, Maria C Renner, Martina Moeton, Marco van Eijk, Rolf G Boot, Willem Kamphuis, Elly M Hol, Jan Aten, Hermen S Overkleeft, Andries Kalsbeek, Johannes M F G Aerts

Netherlands Institute for Neuroscience (NIN)

Research output: Contribution to journal/periodical › Article › Scientific › peer-review

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Abstract

Gaucher disease is characterized by lysosomal accumulation of glucosylceramide due to deficient activity of lysosomal glucocerebrosidase (GBA). In cells, glucosylceramide is also degraded outside lysosomes by the enzyme glucosylceramidase 2 (GBA2) of which inherited deficiency is associated with ataxias. The interest in GBA and glucosylceramide metabolism in the brain has grown following the notion that mutations in the GBA gene impose a risk factor for motor disorders such as α-synucleinopathies. We earlier developed a β-glucopyranosyl-configured cyclophellitol-epoxide type activity based probe (ABP) allowing in vivo and in vitro visualization of active molecules of GBA with high spatial resolution. Labeling occurs through covalent linkage of the ABP to the catalytic nucleophile residue in the enzyme pocket. Here, we describe a method to visualize active GBA molecules in rat brain slices using in vivo labeling. Brain areas related to motor control, like the basal ganglia and motor related structures in the brainstem, show a high content of active GBA. We also developed a β-glucopyranosyl cyclophellitol-aziridine ABP allowing in situ labeling of GBA2. Labeled GBA2 in brain areas can be identified and quantified upon gel electrophoresis. The distribution of active GBA2 markedly differs from that of GBA, being highest in the cerebellar cortex. The histological findings with ABP labeling were confirmed by biochemical analysis of isolated brain areas. In conclusion, ABPs offer sensitive tools to visualize active GBA and to study the distribution of GBA2 in the brain and thus may find application to establish the role of these enzymes in neurodegenerative disease conditions such as α-synucleinopathies and cerebellar ataxia.

Original language	English
Pages (from-to)	e0138107
Journal	PLoS One
Volume	10
Issue number	9
DOIs	https://doi.org/10.1371/journal.pone.0138107
Publication status	Published - 2015

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Herrera Moro Chao, D., Kallemeijn, W. W., Marques, A. R. A., Orre, M., Ottenhoff, R., van Roomen, C., Foppen, E., Renner, M. C., Moeton, M., van Eijk, M., Boot, R. G., Kamphuis, W., Hol, E. M., Aten, J., Overkleeft, H. S., Kalsbeek, A., & Aerts, J. M. F. G. (2015). Visualization of Active Glucocerebrosidase in Rodent Brain with High Spatial Resolution following In Situ Labeling with Fluorescent Activity Based Probes. PLoS One, 10(9), e0138107. https://doi.org/10.1371/journal.pone.0138107

@article{cbb61e57b477438db753442d4c9032a4,

title = "Visualization of Active Glucocerebrosidase in Rodent Brain with High Spatial Resolution following In Situ Labeling with Fluorescent Activity Based Probes",

abstract = "Gaucher disease is characterized by lysosomal accumulation of glucosylceramide due to deficient activity of lysosomal glucocerebrosidase (GBA). In cells, glucosylceramide is also degraded outside lysosomes by the enzyme glucosylceramidase 2 (GBA2) of which inherited deficiency is associated with ataxias. The interest in GBA and glucosylceramide metabolism in the brain has grown following the notion that mutations in the GBA gene impose a risk factor for motor disorders such as α-synucleinopathies. We earlier developed a β-glucopyranosyl-configured cyclophellitol-epoxide type activity based probe (ABP) allowing in vivo and in vitro visualization of active molecules of GBA with high spatial resolution. Labeling occurs through covalent linkage of the ABP to the catalytic nucleophile residue in the enzyme pocket. Here, we describe a method to visualize active GBA molecules in rat brain slices using in vivo labeling. Brain areas related to motor control, like the basal ganglia and motor related structures in the brainstem, show a high content of active GBA. We also developed a β-glucopyranosyl cyclophellitol-aziridine ABP allowing in situ labeling of GBA2. Labeled GBA2 in brain areas can be identified and quantified upon gel electrophoresis. The distribution of active GBA2 markedly differs from that of GBA, being highest in the cerebellar cortex. The histological findings with ABP labeling were confirmed by biochemical analysis of isolated brain areas. In conclusion, ABPs offer sensitive tools to visualize active GBA and to study the distribution of GBA2 in the brain and thus may find application to establish the role of these enzymes in neurodegenerative disease conditions such as α-synucleinopathies and cerebellar ataxia.",

author = "{Herrera Moro Chao}, Daniela and Kallemeijn, {Wouter W} and Marques, {Andre R A} and Marie Orre and Roelof Ottenhoff and {van Roomen}, Cindy and Ewout Foppen and Renner, {Maria C} and Martina Moeton and {van Eijk}, Marco and Boot, {Rolf G} and Willem Kamphuis and Hol, {Elly M} and Jan Aten and Overkleeft, {Hermen S} and Andries Kalsbeek and Aerts, {Johannes M F G}",

year = "2015",

doi = "10.1371/journal.pone.0138107",

language = "English",

volume = "10",

pages = "e0138107",

journal = "PLoS One",

issn = "1932-6203",

publisher = "Public Library of Science",

number = "9",

}

Herrera Moro Chao, D, Kallemeijn, WW, Marques, ARA, Orre, M, Ottenhoff, R, van Roomen, C, Foppen, E, Renner, MC, Moeton, M, van Eijk, M, Boot, RG, Kamphuis, W, Hol, EM, Aten, J, Overkleeft, HS, Kalsbeek, A & Aerts, JMFG 2015, 'Visualization of Active Glucocerebrosidase in Rodent Brain with High Spatial Resolution following In Situ Labeling with Fluorescent Activity Based Probes', PLoS One, vol. 10, no. 9, pp. e0138107. https://doi.org/10.1371/journal.pone.0138107

Visualization of Active Glucocerebrosidase in Rodent Brain with High Spatial Resolution following In Situ Labeling with Fluorescent Activity Based Probes. / Herrera Moro Chao, Daniela; Kallemeijn, Wouter W; Marques, Andre R A et al.

In: PLoS One, Vol. 10, No. 9, 2015, p. e0138107.

Research output: Contribution to journal/periodical › Article › Scientific › peer-review

TY - JOUR

T1 - Visualization of Active Glucocerebrosidase in Rodent Brain with High Spatial Resolution following In Situ Labeling with Fluorescent Activity Based Probes

AU - Herrera Moro Chao, Daniela

AU - Kallemeijn, Wouter W

AU - Marques, Andre R A

AU - Orre, Marie

AU - Ottenhoff, Roelof

AU - van Roomen, Cindy

AU - Foppen, Ewout

AU - Renner, Maria C

AU - Moeton, Martina

AU - van Eijk, Marco

AU - Boot, Rolf G

AU - Kamphuis, Willem

AU - Hol, Elly M

AU - Aten, Jan

AU - Overkleeft, Hermen S

AU - Kalsbeek, Andries

AU - Aerts, Johannes M F G

PY - 2015

Y1 - 2015

N2 - Gaucher disease is characterized by lysosomal accumulation of glucosylceramide due to deficient activity of lysosomal glucocerebrosidase (GBA). In cells, glucosylceramide is also degraded outside lysosomes by the enzyme glucosylceramidase 2 (GBA2) of which inherited deficiency is associated with ataxias. The interest in GBA and glucosylceramide metabolism in the brain has grown following the notion that mutations in the GBA gene impose a risk factor for motor disorders such as α-synucleinopathies. We earlier developed a β-glucopyranosyl-configured cyclophellitol-epoxide type activity based probe (ABP) allowing in vivo and in vitro visualization of active molecules of GBA with high spatial resolution. Labeling occurs through covalent linkage of the ABP to the catalytic nucleophile residue in the enzyme pocket. Here, we describe a method to visualize active GBA molecules in rat brain slices using in vivo labeling. Brain areas related to motor control, like the basal ganglia and motor related structures in the brainstem, show a high content of active GBA. We also developed a β-glucopyranosyl cyclophellitol-aziridine ABP allowing in situ labeling of GBA2. Labeled GBA2 in brain areas can be identified and quantified upon gel electrophoresis. The distribution of active GBA2 markedly differs from that of GBA, being highest in the cerebellar cortex. The histological findings with ABP labeling were confirmed by biochemical analysis of isolated brain areas. In conclusion, ABPs offer sensitive tools to visualize active GBA and to study the distribution of GBA2 in the brain and thus may find application to establish the role of these enzymes in neurodegenerative disease conditions such as α-synucleinopathies and cerebellar ataxia.

AB - Gaucher disease is characterized by lysosomal accumulation of glucosylceramide due to deficient activity of lysosomal glucocerebrosidase (GBA). In cells, glucosylceramide is also degraded outside lysosomes by the enzyme glucosylceramidase 2 (GBA2) of which inherited deficiency is associated with ataxias. The interest in GBA and glucosylceramide metabolism in the brain has grown following the notion that mutations in the GBA gene impose a risk factor for motor disorders such as α-synucleinopathies. We earlier developed a β-glucopyranosyl-configured cyclophellitol-epoxide type activity based probe (ABP) allowing in vivo and in vitro visualization of active molecules of GBA with high spatial resolution. Labeling occurs through covalent linkage of the ABP to the catalytic nucleophile residue in the enzyme pocket. Here, we describe a method to visualize active GBA molecules in rat brain slices using in vivo labeling. Brain areas related to motor control, like the basal ganglia and motor related structures in the brainstem, show a high content of active GBA. We also developed a β-glucopyranosyl cyclophellitol-aziridine ABP allowing in situ labeling of GBA2. Labeled GBA2 in brain areas can be identified and quantified upon gel electrophoresis. The distribution of active GBA2 markedly differs from that of GBA, being highest in the cerebellar cortex. The histological findings with ABP labeling were confirmed by biochemical analysis of isolated brain areas. In conclusion, ABPs offer sensitive tools to visualize active GBA and to study the distribution of GBA2 in the brain and thus may find application to establish the role of these enzymes in neurodegenerative disease conditions such as α-synucleinopathies and cerebellar ataxia.

U2 - 10.1371/journal.pone.0138107

DO - 10.1371/journal.pone.0138107

M3 - Article

C2 - 26418157

SN - 1932-6203

VL - 10

SP - e0138107

JO - PLoS One

JF - PLoS One

IS - 9

ER -

Visualization of Active Glucocerebrosidase in Rodent Brain with High Spatial Resolution following In Situ Labeling with Fluorescent Activity Based Probes

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