NOVEL GLYCOCONJUGATE-BINDING PROTEINS OF LEUCOCYTES

膜相关的碳水化合物结合分子似乎 参与各种生理上有意义的互动。 Galaptins，一类内源性保守的，β- 半乳糖苷结合凝集素似乎在生长中发挥作用 控制、细胞黏附和转移现象与免疫 调制。Galaptin似乎在人体组织中普遍存在 但在人类白细胞中有差异表达。它是 该项目的目的是执行物理-和 银杏叶提取物Galaptin的免疫化学特性 人类的棕黄色被毛细胞。Galaptin将与正常分离 人外周血白细胞去唾液酸-琼脂糖亲和试验 层析法。纯度、相对分子质量和亚基 成分将通过单层和二维电泳法进行分析 通过免疫印迹和免疫检测。 用FPLC凝胶过滤对天然Galaptin进行分析。物理上- Galaptin的化学特性将包括氨基酸 组成和氨基酸序列分析。构图 将对Waters Pico-Tag系统进行研究。 将对Galaptin的降解进行序列研究 用应用生物系统470A测序仪测序。这个 将确定手用抗Galaptin兔血清的特异性。 Galaptin碳水化合物结合特异性将被表征 利用酶联免疫吸附试验和亲和珠法。结合部位 探针将包括表征良好的糖偶联物和它们的 降解产品以及一个完全 描述了合成碳水化合物的特征。这个 Galaptin在白细胞亚型中的分布将是 由免疫组织化学程序确定使用我们的抗- Galaptin血清。本申请中描述的研究将引导 与碳水化合物结合的物理化学特征有关 人外周血白细胞中存在的蛋白质及其鉴定 起源的细胞。这些研究将奠定必要的基础 进一步阐明糖类结合蛋白的功能 以及功能的干扰如何影响细胞的行为。

Membrane-associated carbohydrate-binding molecules appear to be involved in a variety of physiologically significant interactions. Galaptins, a class of endogenous well-conserved, beta- galactoside-binding lectins, appear to play a role in growth control, cell adhesion and metastatic phenomena and immune modulation. Galaptin appears to be ubiquitous in human tissues but is differentially expressed in human leucocytes. It is the purpose of this project to carry out the physico- and immunochemical characterization of galaptin isolated from human buffy coat cells. Galaptin will be isolated from normal human peripheral leucocytes by asialofetuin-Sepharose affinity chromatography. The purity, molecular weight and subunit composition will be analyzed by single and 2-D electrophoretic procedures and by Western blotting followed by immunodetection. Native galaptin will be analyzed by FPLC gel filtration. Physico- chemical characterization of galaptin will include amino acid composition and amino acid sequence analyses. Compositional studies will be carried out on a Waters Pico-Tag system. Sequence studies will be carried out on galaptin degradation fragments with an Applied Biosystems 470A sequenator. The specificity of anti-galaptin rabbit sera in hand will be determined. Galaptin carbohydrate-binding specificity will be characterize utilizing ELISA and affinity bead methodologies. Binding-site probes will include well-characterize glycoconjugates and their degradation products as well as a large battery of completely characterized synthetic carbohydrate compounds. The distribution of galaptin among leucocyte subtypes will be determined by immunohistochemical procedures utilizing our anti- galaptin sera. The research described in this application will lead to the physico-chemical characterization of a carbohydrate-biding protein present in human peripheral leucocytes and identification of cells of origin. These studies will lay the necessary foundation to pursue the elucidation of carbohydrate-binding protein function and how pertubation of function affects cell behavior.