节点文献
基质辅助激光解吸电离飞行时间质谱法区分大肠杆菌糖基转移酶基因
Differentiation of Specific Function of Glycosyltransferase Gene by Matrix Assisted Laser Desorption-Time of FIight-Mass Spectrometry
【摘要】 采用高能碰撞诱导解离(CID)-负离子模式基质辅助激光解吸离子化-飞行时间质谱技术(MALDITOF MS)区别wfgD(大肠杆菌O152中O抗原基因簇内)编码的β-1,3-葡萄糖转移酶和wfgD(大肠杆菌O77中O抗原基因簇内)编码的α-1,3-甘露糖转移酶酶促反应产物-两个脂寡糖非对应异构体。结果表明:由高能CID-MALDI TOF质谱图中产物离子[B3]/[Y3]丰度比率的差别可明确区别两个酶促反应产物。并发现碰撞能量为1.0kV时,碰撞靶气的压力变化可影响产物离子[B3]/[Y3]的丰度比率。推断MALDI-TOF质谱图中,两个脂寡糖非对应异构体的磷酸二脂键部分和糖苷键的裂解依赖于糖基给予体的立体化学。
【Abstract】 The glycosyltransferases(GTs) assembling the O-chain utilize lipid-linked acceptor substrates and nucleotide sugar donor substrates.The difficulty in characterization and discriminate enzymatic products preclude the elucidation of the specific function of putative GTs.In this study,high-energy collision induced dissociation(CID)-negative-ion mode matrix-assisted laser desorption-ionization time-of-flight mass spectrometry(MALDI TOF MS) was used to differentiatewfg D(in E.coli O152 O antigen gene cluste) product andwfg D(in E.coli O77 O antigen gene cluste) product.By the mass number analysis,it was found that nearly all of the CID fragment ions were produced by cleavages of diphosphate moiety and glycosidic linkage,while no ring fragmentation was produced.The experimental results demonstrate that the differences in the intensity ratio of fragment ions of [B3]/[Y3]allowed unambiguous differentiation ofwfg D product andwfg D product,and when 1 kV is used as the collision energy,the change of collision gas pressure can affect the intensity ratio of fragment ions of [B3]/[Y3].It was concluded that the abundances of CID fragment ions depended on the stereochemistry of the glycosyl donors in the cleavage of the diphosphate moiety and glycosidic linkage of the lipooligosaccharides(LOS) enzymatic product in MALDI-TOF mass spectra,and CID-MALDI TOF MS analysis of LOS enzymatic product is likely to prove an invaluable tool that can be used to determine the pecificity of Oantigen GT genes.
【Key words】 Glucosyltransferase; Lipooligosaccharide; Matrix-assisted laser desorption-time of flight-mass spectrometry;
- 【文献出处】 分析化学 ,Chinese Journal of Analytical Chemistry , 编辑部邮箱 ,2011年01期
- 【分类号】R378
- 【被引频次】1
- 【下载频次】211