KINETIC ANALYSIS OF TOXIN-RECEPTOR INTERACTIONS

这个子项目是许多利用资源的研究子项目之一 由NIH/NCRR资助的中心拨款提供。子项目的主要支持 而子项目的主要调查员可能是由其他来源提供的， 包括其它NIH来源。 列出的子项目总成本可能 代表子项目使用的中心基础设施的估计数量， 而不是由NCRR赠款提供给子项目或子项目工作人员的直接资金。 许多重要的生物反应发生在双层膜表面之内或之上。 这类反应的一个重要类别是可溶性配体与其细胞表面受体之间的相互作用。 配体-受体相互作用可以是单价的，如小肽配体的情况，二价的，如许多大肽激素和抗体的情况，或甚至更高价的。 后一类是许多细菌毒素与其细胞表面受体之间的相互作用。 霍乱毒素进入哺乳动物细胞，例如通过五聚体B亚基与细胞膜中的神经节苷脂Gm 1的结合来介导。 这种相互作用的显着特征包括单个B亚基与Gm 1的碳水化合物部分之间的低亲和力单价相互作用、五聚体B亚基与几个膜结合的Gm 1分子之间的高亲和力多价相互作用，以及Gm 1分子在双层膜内的横向移动性。 我们最近将我们对固定化双分析剂的研究扩展到活细胞中毒素加工的研究。 使用动力学和温度控制的流式细胞术，我们正在研究的速率和能力的各种内化途径在活细胞。 这些结果被用于建立和测试细胞内配体加工模型。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Many critical biological reactions occur in or on bilayer membrane surfaces. An important class of such reactions is the interactions between soluble ligands and their cell-surface receptors. Ligand-receptor interactions may be monovalent, as is the case for small peptide ligands, bivalent, as for many large peptide hormones and antibodies, or of even higher valencies. In the latter class are the interactions between many bacterial toxins and their cell-surface receptors. Cholera toxin entry into mammalian cells, for example is mediated by binding of the pentameric B subunit to ganglioside Gm1 in the cell membrane. Notable features of this interaction include a low affinity monovalent interaction between individual B subunits and the carbohydrate moiety of Gm1, a high affinity multivalent interaction between pentameric B subunit with several membrane-bound Gm1 molecules, and the lateral mobility of the Gm1 molecule within the bilayer membrane. We have recently extended our studies on immobilized bialyers to the study of toxin processing in live cells. Using kinetic and temperature controlled flow cytometry, we are investigating the rates and capacities of various internalization pathways in live cells. These results are being used to build and test models of intracellular ligand processing.