Molecules and Cells

Korean Society for Molecular and Cellular Biology (한국분자세포생물학회)
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Quantitative and Rapid Analysis of Transglutaminase Activity Using Protein Arrays in Mammalian Cells

  • Kwon, Mi-Hye (Department of Molecular and Cellular Biochemistry and Vascular System Research Center, Kangwon National University School of Medicine) ;
  • Jung, Jae-Wan (Department of Molecular and Cellular Biochemistry and Vascular System Research Center, Kangwon National University School of Medicine) ;
  • Jung, Se-Hui (Department of Molecular and Cellular Biochemistry and Vascular System Research Center, Kangwon National University School of Medicine) ;
  • Park, Jin-Young (Department of Molecular and Cellular Biochemistry and Vascular System Research Center, Kangwon National University School of Medicine) ;
  • Kim, Young-Myeong (Department of Molecular and Cellular Biochemistry and Vascular System Research Center, Kangwon National University School of Medicine) ;
  • Ha, Kwon-Soo (Department of Molecular and Cellular Biochemistry and Vascular System Research Center, Kangwon National University School of Medicine)
  • Received : 2008.10.21
  • Accepted : 2009.01.07
  • Published : 2009.03.31
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Abstract

We developed a novel on-chip activity assay using protein arrays for quantitative and rapid analysis of transglutaminase activity in mammalian cells. Transglutaminases are a family of $Ca^{2+}$-dependent enzymes involved in cell regulation as well as human diseases such as neurodegenerative disorders, inflammatory diseases and tumor progression. We fabricated the protein arrays by immobilizing N,N'-dimethylcasein (a substrate) on the amine surface of the arrays. We initiated transamidating reaction on the protein arrays and determined the transglutaminase activity by analyzing the fluorescence intensity of biotinylated casein. The on-chip transglutaminase activity assay was proved to be much more sensitive than the $[^3H]putrescine$-incorporation assay. We successfully applied the on-chip assay to a rapid and quantitative analysis of the transglutaminase activity in all-trans retinoic acid-treated NIH 3T3 and SH-SY5Y cells. In addition, the on-chip transglutaminase activity assay was sufficiently sensitive to determine the transglutaminase activity in eleven mammalian cell lines. Thus, this novel on-chip transglutaminase activity assay was confirmed to be a sensitive and high-throughput approach to investigating the roles of transglutaminase in cellular signaling, and, moreover, it is likely to have a strong potential for monitoring human diseases.

Keywords

Acknowledgement

Supported by : Ministry of Health, Welfare and Family Affairs

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