Fluorescence-Quenched Substrates for Quantitative Live Cell Imaging of Glucocerebrosidase Activity
Glucocerebrosidase (GCase) is a lysosomal glycoside hydrolase that cleaves the glycolipid glucosylceramide (GlcCer). Deficiencies of this enzyme lead to accumulation of GlcCer and the development of the lysosomal storage disease known as Gaucher's disease. Recently, loss-of-function mutations i...
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| Veröffentlicht in: | Methods in enzymology 2018, Vol.598, p.199-215 |
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| creator | Ashmus, Roger A Shen, David L Vocadlo, David J |
| description | Glucocerebrosidase (GCase) is a lysosomal glycoside hydrolase that cleaves the glycolipid glucosylceramide (GlcCer). Deficiencies of this enzyme lead to accumulation of GlcCer and the development of the lysosomal storage disease known as Gaucher's disease. Recently, loss-of-function mutations in the GBA1 gene that encodes GCase have been linked to Parkinson's disease. Currently pursued therapeutic strategies to increase GCase involve enzyme replacement therapy, chemical chaperone therapy, and GCase activators. A challenge associated with advancing such strategies is to efficiently monitor GCase activity within the lysosomes of live cells. In this chapter, we review the design and use of the fluorescent-quenched probe GBA1-FQ2 to quantitatively measure GCase activity in lysosomes of live cells. |
| doi_str_mv | 10.1016/bs.mie.2017.06.014 |
| format | Article |
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Deficiencies of this enzyme lead to accumulation of GlcCer and the development of the lysosomal storage disease known as Gaucher's disease. Recently, loss-of-function mutations in the GBA1 gene that encodes GCase have been linked to Parkinson's disease. Currently pursued therapeutic strategies to increase GCase involve enzyme replacement therapy, chemical chaperone therapy, and GCase activators. A challenge associated with advancing such strategies is to efficiently monitor GCase activity within the lysosomes of live cells. In this chapter, we review the design and use of the fluorescent-quenched probe GBA1-FQ2 to quantitatively measure GCase activity in lysosomes of live cells.</description><identifier>EISSN: 1557-7988</identifier><identifier>DOI: 10.1016/bs.mie.2017.06.014</identifier><identifier>PMID: 29306435</identifier><language>eng</language><publisher>United States</publisher><subject>Cells, Cultured ; Enzyme Assays - instrumentation ; Enzyme Assays - methods ; Fibroblasts ; Fluorescence ; Fluorescent Dyes - chemistry ; Glucosylceramidase - analysis ; Glucosylceramidase - metabolism ; Humans ; Intravital Microscopy - instrumentation ; Intravital Microscopy - methods ; Lysosomes - metabolism ; Microscopy, Fluorescence - instrumentation ; Microscopy, Fluorescence - methods ; Primary Cell Culture - instrumentation ; Primary Cell Culture - methods ; Spectrometry, Fluorescence - instrumentation ; Spectrometry, Fluorescence - methods</subject><ispartof>Methods in enzymology, 2018, Vol.598, p.199-215</ispartof><rights>2018 Elsevier Inc. 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| ispartof | Methods in enzymology, 2018, Vol.598, p.199-215 |
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| subjects | Cells, Cultured Enzyme Assays - instrumentation Enzyme Assays - methods Fibroblasts Fluorescence Fluorescent Dyes - chemistry Glucosylceramidase - analysis Glucosylceramidase - metabolism Humans Intravital Microscopy - instrumentation Intravital Microscopy - methods Lysosomes - metabolism Microscopy, Fluorescence - instrumentation Microscopy, Fluorescence - methods Primary Cell Culture - instrumentation Primary Cell Culture - methods Spectrometry, Fluorescence - instrumentation Spectrometry, Fluorescence - methods |
| title | Fluorescence-Quenched Substrates for Quantitative Live Cell Imaging of Glucocerebrosidase Activity |
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