Ito Hiromi, Chiba Yasunori, Kameyama Akihiko, Sato Takashi, Narimatsu Hisashi
Research Center for Medical Glycoscience, National Institute of Advanced Industrial Science and Technology (AIST), Open Space Laboratory C-2, Umezono, Tsukuba, Ibaraki, Japan.
Methods Enzymol. 2010;478:127-49. doi: 10.1016/S0076-6879(10)78005-8.
In the GlycoGene Project, we have comprehensively cloned novel human genes associated with the synthesis of glycans using bioinformatics technology. Recombinant glycosyltransferases can be expressed in various expression systems. Diverse glycan structures are easily and rapidly achieved using these glycosyltransferases in vitro. Additionally, we have developed an in vivo production system for mammalian mucin-type glycopeptides using a genetically engineered yeast strain. This system enables the generation of glycopeptides which are O-glycosylated on a specific position by introducing different types of ppGalNAc-T genes. As an application of the glycan and glycopeptide libraries to glycan analysis, we have constructed a multistage tandem mass (MS(n)) spectral database containing observed MS(n) spectra. Using the MS(n) spectral database, it is possible to identify glycan structures very easily and rapidly by spectral matching.
在糖基因项目中，我们利用生物信息学技术全面克隆了与聚糖合成相关的新型人类基因。重组糖基转移酶可在多种表达系统中表达。使用这些糖基转移酶可在体外轻松快速地实现多种聚糖结构。此外，我们利用基因工程酵母菌株开发了一种用于哺乳动物粘蛋白型糖肽的体内生产系统。通过引入不同类型的ppGalNAc-T基因，该系统能够生成在特定位置进行O-糖基化的糖肽。作为聚糖和糖肽文库在聚糖分析中的应用，我们构建了一个包含观察到的多级串联质谱（MS(n)）光谱的光谱数据库。利用该MS(n)光谱数据库，通过光谱匹配可以非常轻松快速地鉴定聚糖结构。
