Force Probe Investigations of NCAM Adhesion

描述（由申请人提供）：我们的目标是通过 直接力测量和基因操作的组合， 蛋白质结构，嗜同性粘附的基本物理机制 神经细胞粘附分子（NCAM）与NCAM调节之间的关系 通过翻译后修饰的粘附。NCAM是一个大型的多域 通过嗜同性结合介导细胞间相互作用的蛋白质。是 一种最丰富和广泛的细胞粘附蛋白， 免疫球蛋白超家族，分子的粘附功能已被证明 来影响一些重要的发育过程。述蛋白质是 不寻常的是，它表现出两种结构形式：其中一种表现出 粘附活性和另一种显示抗粘附功能。虽然 广泛的研究解决了这些功能的生物学作用， 已经在分子细节中定义了潜在的机制。的 抗粘附性与蛋白质糖基化有关， 聚唾液酸（PSA）。这种修饰不仅影响NCAM粘附， 以及由其它粘附分子介导的细胞相互作用。这些拟议 研究将量化膜之间力的距离依赖性 显示不同形式的NCAM，以确定分子 聚唾液酸改变NCAM之间相互作用的机制， 以及其他两种粘附蛋白N-cadherin和Li的相互作用。的 测量将直接检验PSA在空间上阻碍细胞增殖的假设。 细胞的紧密贴壁，从而对 细胞相互作用关于粘合剂功能，附加的 NCAM和NCAM碎片均缺乏特定的力/距离测量 域将进一步测试目前的几个假设， 结构域相互作用介导的亲同性结合。这些拟议 力距测量将产生独特的分子水平信息 关于NCAM粘附的机制， 全细胞研究、溶液结合测量或晶体结构。

DESCRIPTION (provided by applicant): Our objectives are to identify, by a combination of direct force measurements and the genetic manipulation of protein structure, the fundamental physical mechanisms of homophilic adhesion between neural cell adhesion molecules (NCAM) and the regulation of NCAM adhesion by post-translational modification. NCAM is a large, multidomain protein that mediates cell-cell interactions through homophilic binding. It is one of the most abundant and widespread cell adhesion proteins of the immunoglobulin superfamily, and the molecule's adhesive function has been shown to influence a number of important developmental processes. The protein is unusual in that it exhibits two structural forms: one of which exhibits adhesive activity and the other which displays anti-adhesive function. Although extensive research addressed the biological role of these functions, neither of the underlying mechanisms has been defined in molecular detail. The anti-adhesive property is linked to the protein glycosylation, which produces polysialylic acid (PSA). This modification affects not only NCAM adhesion but also cell interactions mediated by other adhesion molecules. These proposed studies will quantify the distance dependence of the forces between membranes displaying the different forms of NCAM, in order to determine the molecular mechanism by which polysialic acid alters both interactions between NCAM as well as the interactions of two other adhesion proteins N-cadherin and Li. The measurements will directly test the hypothesis that PSA sterically impedes the close apposition of cells and thereby exerts a general regulatory influence on cell interactions. With regard to the adhesive function, additional force/distance measurements with both NCAM and NCAM fragments lacking specific domains will further test several current hypotheses regarding the specific domain interactions that mediate homophilic binding. These proposed force-distance measurements will generate unique, molecular level information concerning the mechanisms of NCAM adhesion that cannot be determined from studies with whole cells, solution binding measurements, or crystal structures.