Abstract
The majority of eukaryotic proteins undergo post-translational modifications (PTMs) involving the attachment of complex glycans, predominantly through N-glycosylation and O-glycosylation. PTMs play important roles in virtually all cellular processes, and aberrant regulation of protein glycosylation and glycan processing has been implicated in various diseases. However, glycan processing on proteins in various cellular contexts has not been visualized. We had previously developed a quinone methide cleavage (QMC) platform for enhanced substrate design. This platform was applied here to screen for novel glycan-processing enzymes. We designed and synthesized fluorescent substrates with β-allopyranoside residues using the QMC platform. When applied in cell-based assays, the fluorescent substrates allowed rapid and clear visualization of β-allosidase activity in the Golgi apparatus of human cultured cells. The QMC platform will likely find broad applications in visualizing the activities of glycan processing enzymes in living cells and in studying PTMs.
| Original language | English |
|---|---|
| Pages (from-to) | 73-79 |
| Number of pages | 7 |
| Journal | Bioorganic and Medicinal Chemistry |
| Volume | 23 |
| Issue number | 1 |
| DOIs | |
| Publication status | Published - 2015 Jan 1 |
| Externally published | Yes |
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Keywords
- A EM emission wavelength
- Abbreviations 2MeTG 2-methyl
- BFA brefeldin
- ER endoplasmic reticulum
- EX excitation wavelength Man mannose
- MS mass spectrometry
- NMR nuclear magnetic resonance
- PBS phosphate-buffered saline
- PTM post-translational modifications
- QMC quinone methide cleavage
- TBP tributyl phosphine
- TFMU 4-Trifluoromethylumbelliferone
- TokyoGreen
ASJC Scopus subject areas
- Biochemistry
- Molecular Medicine
- Molecular Biology
- Pharmaceutical Science
- Drug Discovery
- Clinical Biochemistry
- Organic Chemistry
Cite this
Identification of a novel glycan processing enzyme with exo-acting β-allosidase activity in the Golgi apparatus using a new platform for the synthesis of fluorescent substrates. / Hakamata, Wataru; Miura, Kazuki; Hirano, Takako; Nishio, Toshiyuki.
In: Bioorganic and Medicinal Chemistry, Vol. 23, No. 1, 01.01.2015, p. 73-79.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Identification of a novel glycan processing enzyme with exo-acting β-allosidase activity in the Golgi apparatus using a new platform for the synthesis of fluorescent substrates
AU - Hakamata, Wataru
AU - Miura, Kazuki
AU - Hirano, Takako
AU - Nishio, Toshiyuki
PY - 2015/1/1
Y1 - 2015/1/1
N2 - The majority of eukaryotic proteins undergo post-translational modifications (PTMs) involving the attachment of complex glycans, predominantly through N-glycosylation and O-glycosylation. PTMs play important roles in virtually all cellular processes, and aberrant regulation of protein glycosylation and glycan processing has been implicated in various diseases. However, glycan processing on proteins in various cellular contexts has not been visualized. We had previously developed a quinone methide cleavage (QMC) platform for enhanced substrate design. This platform was applied here to screen for novel glycan-processing enzymes. We designed and synthesized fluorescent substrates with β-allopyranoside residues using the QMC platform. When applied in cell-based assays, the fluorescent substrates allowed rapid and clear visualization of β-allosidase activity in the Golgi apparatus of human cultured cells. The QMC platform will likely find broad applications in visualizing the activities of glycan processing enzymes in living cells and in studying PTMs.
AB - The majority of eukaryotic proteins undergo post-translational modifications (PTMs) involving the attachment of complex glycans, predominantly through N-glycosylation and O-glycosylation. PTMs play important roles in virtually all cellular processes, and aberrant regulation of protein glycosylation and glycan processing has been implicated in various diseases. However, glycan processing on proteins in various cellular contexts has not been visualized. We had previously developed a quinone methide cleavage (QMC) platform for enhanced substrate design. This platform was applied here to screen for novel glycan-processing enzymes. We designed and synthesized fluorescent substrates with β-allopyranoside residues using the QMC platform. When applied in cell-based assays, the fluorescent substrates allowed rapid and clear visualization of β-allosidase activity in the Golgi apparatus of human cultured cells. The QMC platform will likely find broad applications in visualizing the activities of glycan processing enzymes in living cells and in studying PTMs.
KW - A EM emission wavelength
KW - Abbreviations 2MeTG 2-methyl
KW - BFA brefeldin
KW - ER endoplasmic reticulum
KW - EX excitation wavelength Man mannose
KW - MS mass spectrometry
KW - NMR nuclear magnetic resonance
KW - PBS phosphate-buffered saline
KW - PTM post-translational modifications
KW - QMC quinone methide cleavage
KW - TBP tributyl phosphine
KW - TFMU 4-Trifluoromethylumbelliferone
KW - TokyoGreen
UR - http://www.scopus.com/inward/record.url?scp=84918820034&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=84918820034&partnerID=8YFLogxK
U2 - 10.1016/j.bmc.2014.11.023
DO - 10.1016/j.bmc.2014.11.023
M3 - Article
C2 - 25497961
AN - SCOPUS:84918820034
VL - 23
SP - 73
EP - 79
JO - Bioorganic and Medicinal Chemistry
JF - Bioorganic and Medicinal Chemistry
SN - 0968-0896
IS - 1
ER -