Glycosyl conjugates of biotinylated diaminopyridine applied for study of carbohydrate-to-carbohydrate interaction.
Glycosyl conjugates of biotinylated diaminopyridine applied for study of carbohydrate-to-carbohydrate interaction.
复制标题
生物素化二氨基吡啶的糖基缀合物应用于研究碳水化合物与碳水化合物的相互作用。
DOI：
10.1007/s10719-010-9304-3
复制
发表时间：
2010
影响因子：
3
通讯作者：
Hakomori,Sen-Itiroh
中科院分区：
文献类型：
--
作者：
Utkina,Natalia;Yoon,Seon-Joo;Hakomori,Sen-Itiroh
我们和其他人之前的研究表明，细胞间粘附是由特定的碳水化合物与碳水化合物相互作用（CCI）介导的。先前的研究基于各种生物化学和生物物理方法，包括使用带有荧光标签的标记糖基表位。然而，这些方法理想地要求糖基表位必须固定至固相分子，优选以多价固定。本研究的目的是使用与生物素化二氨基吡啶 (BAP) 缀合的特定糖基残基建立 CCI 过程，并观察：(i) 具有 5-6 个 GlcNAc 末端的“Os Fr.B”与具有 5-6 个 GlcNAc 末端的“Os Fr.B”之间明显存在同型 CCI。具有 2 个 GlcNAc 末端的“Os Fr.1”之间不存在此类同型 CCI； (ii) GM3 神经节苷脂和 Os Fr.B 与 Os Fr.B 之间异型 CCI 的发生GM3 和 Os Fr.1 之间不存在这种异型 CCI。 Os Fr.B-BAP 缀合物和 Os Fr.B-神经酰胺模拟物 (Os Fr.B-mCer) 之间的相互作用基于两个实验得到证明：(i) 在二价阳离子存在的情况下观察到 Os Fr.B-BAP 缀合物与涂有 Os Fr.B-mCer 的聚苯乙烯板的剂量依赖性结合，这是所有 CCI 过程的先决条件，并且这种结合被 EDTA 消除； (ii)等纳摩尔Os Fr.B-BAP和Os Fr.B-mCer之间的结合被没有缀合物的mM浓度Os Fr.B以剂量依赖性方式抑制。因此，使用糖基表位的BAP缀合物清楚地观察到基于具有多个GlcNAc末端的N-连接聚糖之间的同型CCI以及GM3和此类聚糖之间的异型CCI的细胞粘附过程。
Previous studies by us and others established that cell-cell adhesion is mediated by specific carbohydrate-to-carbohydrate interaction (CCI). Those previous studies were based on various biochemical and biophysical approaches, including the use of labeled glycosyl epitopes with fluorescent tag. However, these methods ideally require that the glycosyl epitope must be fixed to a solid phase molecule, preferably with multivalency. The purpose of the present study is to establish a CCI process using specific glycosyl residues conjugated to biotinylated diaminopyridine (BAP), and to observe: (i) clear occurrence of homotypic CCI between “Os Fr.B” having 5–6 GlcNAc termini,vs. absence of such homotypic CCI between “Os Fr.1” having 2 GlcNAc termini; (ii) occurrence of heterotypic CCI between GM3 ganglioside and Os Fr.B,vs. absence of such heterotypic CCI between GM3 and Os Fr.1. Interaction between Os Fr.B-BAP conjugate and Os Fr.B-ceramide mimetic (Os Fr.B-mCer) was demonstrated based on two experiments: (i) dose-dependent binding of Os Fr.B-BAP conjugate to polystyrene plates coated with Os Fr.B-mCer was observed in the presence of bivalent cation, a prerequisite for all CCI processes, and such binding was abolished by EDTA; (ii) binding between equal nanomolar Os Fr.B-BAP and Os Fr.B-mCer was inhibited by mM concentration Os Fr.B without conjugate, in dose-dependent manner. Thus, cell adhesion processes based on homotypic CCI between N-linked glycans having multiple GlcNAc termini, and heterotypic CCI between GM3 and such glycans, were clearly observed using BAP conjugates of glycosyl epitopes.