Identification and characterization of binding proteins with gangliosides expressed in the brain and nervous system

神经节苷脂是一种含有唾液酸的鞘糖脂，是广泛存在于动物细胞膜上的生理分子。神经节苷脂在结构上的微观异质性已得到证实，但其功能却知之甚少，尽管神经节苷脂被认为是细胞生长和分化、信号转导、细胞粘附等功能分子，但我们最近建立了一种改进的方法，通过纯化鞘糖脂免疫小鼠产生单克隆抗体。利用该方法，我们制备了一系列抗鞘糖脂的抗体并进行了表征。这些抗体不仅在基础科学中，而且在临床医学中有许多可能的应用。在本研究中，我们试图从脑中鉴定和表征神经节苷脂结合蛋白。2000年，我们鉴定了一种GT1b结合蛋白，即脑特异性钠依赖性无机磷酸盐转运蛋白（BNPI）。随后，我们试图表征其与神经节苷脂GT 1 B的结合结构域，并鉴定另一种结合蛋白。关于第一个问题，我们还没有成功地确定，可能是由于BNPI基因导入的细胞不能保持存活的事实。关于第二个问题，我们检测到与GD1a和GD1b相关的大分子，表明它们是粘蛋白。目前，我们正在对这些分子进行测定。我们期望结合蛋白的分析能揭示神经节苷脂在脑中的功能。

Gangliosides, sialic acid-containing glycosphingolipids, are physiological molecules expressed ubiquitously in animal plasma membranes. The micro heterogeneity of gangliosides is well established in structure, but little is known about their function, although gangliosides are implicated as functional molecules such as cell growth and differentiation, signal transduction, cell adhesion and so on. We recently established an improved method for the generation of monoclonal antibodies by immunizing mice with purified glycosphingolipids. Using the method, we generated and characterized a numberof series of antibodies against glycosphingolipids. There are a number of possible applications of these antibodies not only in basic science butalso in clinical medicine. In this study, we tried to identify and characterize ganglioside binding proteins from brain. In 2000, we identified a GT1b binding protein as brain-specific sodium-dependent inorganic phosphate cotrnasporter (BNPI). Subsequently, we attempted to characterize its binding domain with gangiioside GT1 b, and to identify another binding proteins. Regarding with the first issue, we have not succeeded in the determination yet, probably due tothe fact that BNPI gene-introduced cells can not keep alive. Regarding with the second issue, we detected large molecules associated with GD1a and GD1b, suggesting that they are mucin. At present, we are working on the determination of these molecules. We expect that the analysis of the binding proteins should reveal the functionp of gangliosides in the brain.