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Biochemical Titration of Glycogen In vitro
Published on: November 24, 2013
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Novel method for detection of glycogen in cells
Alexander V Skurat1, Dyann M Segvich1, Anna A DePaoli-Roach1
1Department of Biochemistry and Molecular Biology, Indiana University School of Medicine, Indianapolis, IN 46202, USA.
Glycobiology
|January 13, 2017
Summary
Researchers developed a novel recombinant protein probe (GYSC) for detecting glycogen deposits. This tool aids in studying glycogen metabolism and diagnosing glycogen storage diseases (GSDs).
Area of Science:
- Biochemistry
- Molecular Biology
- Cell Biology
Background:
- Glycogen serves as an energy reserve in organisms.
- Genetic defects in glycogen metabolism cause glycogen storage diseases (GSDs).
- Visualizing glycogen is crucial for GSD diagnosis and metabolism studies.
Purpose of the Study:
- To develop a novel, renewable probe for detecting glycogen deposits.
- To utilize a recombinant protein with a carbohydrate-binding module (CBM) for glycogen detection.
Main Methods:
- A fusion protein (GYSC) was engineered, combining glutathione S-transferase, a cMyc epitope, and the Stbd1 CBM.
- Enzyme-linked immunosorbent assay (ELISA) was used to assess GYSC binding to glucose polymers.
- Immunofluorescence staining was performed on cultured cells and isolated mouse muscle fibers.
Main Results:
- GYSC demonstrated binding to glycogen, amylopectin, and amylose.
- Immunofluorescence showed GYSC co-localization with glycogen and glycogen synthase.
- GYSC detected glycogen in lysosomes of Pompe disease models and in muscle fibers of Lafora disease models.
Conclusions:
- The GYSC probe is effective for detecting glycogen in various cellular contexts.
- GYSC facilitates the study of normal and pathological glycogen metabolism.
- This novel probe aids in the diagnosis and research of GSDs.
