Fluorescently labeled inhibitor for profiling cytoplasmic peptide : N-glycanase
抄録
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Cytoplasmic peptide:N-glycanase (PNGase) is an enzyme that removes N-glycans from misfolded glycoproteins. The function of cytoplasmic PNGase plays a significant role in the degradation of misfolded glycoproteins, which is critical for cell viability. Recently, we reported that haloacetoamidyl derivatives of high-mannose-type oligosaccharides selectively modify the catalytic cysteine of cytoplasmic PNGase and serve as its specific inhibitor. Interestingly, a drastically simplified chloroacetamidyl chitobiose derivative [(GlcNAc)(2)-ClAc] was also reactive to PNGase. In our work, it was conjugated to a hydrophobic fluorophore in order to render (GlcNAc)(2)-ClAc cells permeable. We demonstrated that this compound [BODIPY-(GlcNAc)(2)-ClAc] specifically binds to cytoplasmic PNGase from budding yeast (Png1). To date, only Z-VAD-fmk is known as an inhibitor of PNGase. BODIPY-(GlcNAc)(2)-ClAc and Z-VAD-fmk share the same binding site on Png1, while BODIPY-(GlcNAc)(2)-ClAc has markedly stronger inhibitory activity. The functional analysis of PNGase using Z-VAD-fmk should be carefully interpreted because of its intrinsic property as a caspase inhibitor. In sharp contrast, chloroacetamidyl chitobiose was not reactive to caspase. In addition, BODIPY-(GlcNAc)(2)-ClAc did not bind either chitobiose-binding lectins or PNGase from other sources. Moreover, fluorescent microscopy clearly showed that BODIPY-(GlcNAc)(2)-ClAc was efficiently introduced into cells. These results suggest that this compound could be an in vivo inhibitor of cytoplasmic PNGase.
identifier:http://glycob.oxfordjournals.org/cgi/content/full/17/10/1070
収録刊行物
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- GLYCOBIOLOGY
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GLYCOBIOLOGY 17 (10), 1070-1076, 2007
OXFORD UNIV PRESS INC
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詳細情報 詳細情報について
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- CRID
- 1050564287772238720
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- NII論文ID
- 120006385865
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- ISSN
- 09596658
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- Web Site
- http://id.nii.ac.jp/1586/00013373/
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- 本文言語コード
- en
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- 資料種別
- journal article
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- データソース種別
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- IRDB
- CiNii Articles
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