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Sialyltransferase Activity Assay for Ganglioside GM3 Synthase

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Glycolipids

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2613))

Abstract

GM3 synthase (GM3S) is a sialyltransferase that transfers sialic acid from CMP-sialic acid to lactosylceramide. This reaction results in formation of ganglioside GM3 and is essential for biosynthesis of its downstream derivatives, which include a- and b-series gangliosides. Here, we describe a method for GM3S enzymatic assay using fluorescence-labeled alkyl lactoside as acceptor substrate, followed by HPLC for separation of enzymatic product. The method allows quantitative assay of GM3S sialyltransferase activity in cultured cells and mouse brain tissues.

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Acknowledgments

This study was supported by JSPS KAKENHI, grant numbers JP21K06531 (K.-i.I.) and Grant-in-Aid for Scientific Research (B) from MEXT (20H03452), Takeda Foundation, and MEXT-Supported Program for the Strategic Research Foundation at Private Universities (S1201031) (J.-i.I.). The authors are grateful to Dr. S. Anderson for English editing of the manuscript.

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Authors and Affiliations

  1. Division of Glycopathology, Institute of Molecular Biomembrane and Glycobiology, Tohoku Medical and Pharmaceutical University, Sendai, Miyagi, Japan

    Kei-ichiro Inamori, Takahiro Nitta & Jin-ichi Inokuchi

  2. Center for Medical Education, Faculty of Medicine, Tohoku Medical and Pharmaceutical University, Sendai, Miyagi, Japan

    Fumi Shishido & Isao Ohno

  3. Department of Biomolecular Science, Faculty of Science, Toho University, Chiba, Japan

    Soichiro Watanabe

  4. Forefront Research Center, Graduate School of Sciences, Osaka University, Osaka, Japan

    Jin-ichi Inokuchi

Authors
  1. Kei-ichiro Inamori
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  2. Takahiro Nitta
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  3. Fumi Shishido
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  4. Soichiro Watanabe
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  5. Isao Ohno
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  6. Jin-ichi Inokuchi
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Corresponding author

Correspondence to Kei-ichiro Inamori .

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Editors and Affiliations

  1. Department of Chemistry, Graduate School of Science, Osaka University, Osaka, Japan

    Kazuya Kabayama

  2. Institute of Molecular Biomembrane and Glycobiology, Tohoku Medical and Pharmaceutical University, Sendai, Japan

    Jin-ichi Inokuchi

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Inamori, Ki., Nitta, T., Shishido, F., Watanabe, S., Ohno, I., Inokuchi, Ji. (2023). Sialyltransferase Activity Assay for Ganglioside GM3 Synthase. In: Kabayama, K., Inokuchi, Ji. (eds) Glycolipids. Methods in Molecular Biology, vol 2613. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2910-9_9

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  • DOI: https://doi.org/10.1007/978-1-0716-2910-9_9

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-2909-3

  • Online ISBN: 978-1-0716-2910-9

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