Detection of Hydroxyproline O-galactoside by LC/MS
采用 LC/MS检测羟脯氨酸-O-半乳糖苷
发布: 2016年01月20日第6卷第2期 DOI: 10.21769/BioProtoc.1710 浏览次数: 6543
评审: Renate WeizbauerHarrie van ErpAnonymous reviewer(s)
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参见作者原研究论文
The authors used this protocol in:
Mar 2015
Abstract
Hydroxyproline (Hyp) O-galactosylation is a plant-specific post-translational modification found in extracellular glycoproteins such as arabinogalactan proteins (AGPs). Hyp O-galactosylation is mediated by Hyp O-galactosyltransferase (HPGT) that catalyzes the transfer of a D-galactopyranosyl residue to the hydroxyl group of Hyp residues of peptides from the sugar donor UDP-α-D-Gal. Here we describe an LC/MS-based method for the detection of Hyp O-galactoside.
Materials and Reagents
- Cotton
- 1 ml Micropipette tip
- Hyp O-galactosylated peptides or proteins
- 0.22 M Ba(OH)2
- 0.32 M sulfuric acid
- 1 M NaOH
- 1 M HCl
- 10% aqueous ammonia
- 80% acetonitrile containing 0.1% formic acid
- 99.9% acetonitrile (HPLC grade) containing 0.1% formic acid
- Water (HPLC grade) containing 0.1% formic acid
Equipment
- Heat block
- Centrifugal evaporator
- BT AG 50W-X8 Resin (100-200 mg resin, H+ form) (Bio-Rad Laboratories, catalog number: 143-5441 )
- Micro centrifuge
- Micro HPLC system (JASCO International Co., model: micro21 LC-01 )
- LCQ Deca XP-plus ESI ion-trap mass spectrometer (Thermo Fisher Scientific)
- TSK-gel Amide-80 (3 μm) column (2.0 x 150 mm) (Tosoh Bioscience LLC, catalog number: 21865 )
Procedure
文章信息
版权信息
© 2016 The Authors; exclusive licensee Bio-protocol LLC.
如何引用
Ogawa-Ohnishi, M. and Matsubayashi, Y. (2016). Detection of Hydroxyproline O-galactoside by LC/MS. Bio-protocol 6(2): e1710. DOI: 10.21769/BioProtoc.1710.
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分类
植物科学 > 植物生物化学 > 糖类
生物化学 > 糖类 > 多糖
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