MOLECULAR GENETIC ANALYSIS OF LYMPHOCYTE FUNCTION

这一重大项目旨在制定新的战略，以 了解支配初始状态的基本分子机制 免疫反应中的识别事件。作为这样的例子 分子识别事件，我们关注的是分子间的相互作用 主要组织相容性复合体(MHC)分子与特异性T细胞 受体或MHC I类分子与自然杀伤分子上的受体 细胞。我们已经开发了大批量生产的模型系统 涉及的分子：MHC分子、T细胞受体和它们的 片段和NK受体。经过表达和纯化后，这些 分子已被用来检测结构、生物物理参数 具有约束力和功能性。过去一年的主要成就 包括生产足够纯度的T细胞受体结构域 和数量，以允许结晶和高分辨率核 磁共振光谱学。X-射线晶体结构 已经解决了Valpha域的问题，并且高分辨率细化是 正在进行中。一种生产活性T细胞受体的策略 结合特定MHC/肽复合体的特异性结合位点具有 已被证明有效地将细胞表面的MHC/肽复合体染色 活细胞，并阻断特定的T细胞激活。这一战略 为了生产这种具有生物活性的工程T细胞受体 细菌现在已经被应用于额外的T细胞受体。是这样的 研究为管理生物多样性的分子规则提供了新的见解 启动免疫反应，也提供了一种看似合理的方法 为了制造分子，并最终制造出能够特异性抑制T细胞的药物 在自身免疫条件下可能失控的细胞。

This major project has been aimed at developing new strategies to understand the basic molecular mechanisms that govern initial recognition events in the immune response. As examples of such molecular recognition events, we have focused on interactions of the major histocompatibility complex (MHC) molecules with specific T cell receptors or of MHC class I molecules with receptors on natural killer cells. We have developed model systems for producing large quantities of the molecules involved: MHC molecules, T cell receptors and their fragments, and NK receptors. After expression and purification, these molecules have been used to examine structure, biophysical parameters of binding, and function. Major accomplishments in the past year include the production of T cell receptor domains of sufficient purity and quantity to allow both crystallization and high resolution nuclear magnetic resonance spectroscopy. The x-ray crystallographic structure of a Valpha domain has been solved, and high resolution refinement is underway. A strategy for production of an active T cell receptor combining site with specificity for a particular MHC/peptide complex has proven effective in staining cell surface MHC/peptide complexes on living cells, and in blocking specific T cell activation. This strategy for producing such biologically active engineered T cell receptors in bacteria has now been applied to additional T cell receptors. Such studies offer new insights into the molecular rules that govern the initiation of the immune response, and also provide a plausible approach to make molecules and ultimately drugs that can specifically inhibit T cells that might run out of control in autoimmune conditions.