| Budget Amount *help |
¥18,590,000 (Direct Cost: ¥14,300,000、Indirect Cost: ¥4,290,000)
Fiscal Year 2014: ¥5,460,000 (Direct Cost: ¥4,200,000、Indirect Cost: ¥1,260,000)
Fiscal Year 2013: ¥6,240,000 (Direct Cost: ¥4,800,000、Indirect Cost: ¥1,440,000)
Fiscal Year 2012: ¥6,890,000 (Direct Cost: ¥5,300,000、Indirect Cost: ¥1,590,000)
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| Outline of Final Research Achievements |
O-GlcNAcylation of nuclear proteins is a unique reaction in several points of view. O-GlcNAcyation levels are modulated in response to cellular signaling and affect protein localization, activity and stability. We screened engineered lectins to monitor O-GlcNAc modification using mammalian cell surface display from randomized lectin library. Furthermore, we expressed a soluble form of β-N-acetylgalactosaminyltransferase in the nucleus to change O-GlcNAc to GalNAc-GlcNAc. A novel lectin specific for GalNAc-GlcNAc was purified from Wisteria japonica seeds. Using these lectins as a probe, we identified more than 80 kinds of O-GlcNAc-modified proteins including 10 kinds of novel proteins from the nuclear and cytosolic fractions. O-GlcNAc-moidifed peptides were purified by lectin affinity chromatography and identified O-GlcNAc-modified residues by mass spectrometry. A lectin-GFP was expressed in the cytoplasm for monitoring O-GlcNAcylation in a live cell during cell activation.
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