MOLECULAR GENETIC ANALYSIS OF LYMPHOCYTE FUNCTION

这个实验室的重点一直是MHC I类分子， 经典地认为是由46kd组成的分子杂二聚体 多态重链，以小鼠H-2K、D和L为例 分子(类似于人类的HLA-A、-B和-C分子)，即 与12kd的单晶轻链非共价组装， β2-微球蛋白。我们在这些研究中的目标是 检查一些特定的生物合成、结构和 I类分子的功能，包括：1)测量 控制MHC I类结合的生化参数 分子对抗原肽的影响；2)β2-微球蛋白的影响 以及其他血清因素对MHC功能和生化结合的影响 I类分子为抗原肽；3)表达、折叠、 A)中产生的I类分子的组装、结构和稳定性 依赖于RNA的体外翻译系统和b)昆虫病毒/飞蛾 卵巢细胞表达系统；4)表达折叠、组装、 第I类结构域和亚结构域片段的结构和稳定性 重组DNA方法产生的分子，并在组织中表达 培养细胞；5)可溶性MHC-I类分子在 转基因小鼠模型系统；6)单链MHC的制备 第I类分子考察重分子的作用 链/β2-微球蛋白相互作用在细胞功能和稳定性中的作用 这些分子并作为高水平生产的基础 原核生物系统。

The focus of this laboratory has been the MHC class I molecule, classically considered a molecular heterodimer consisting of a 46 kd polymorphic heavy chain, exemplified by the murine H-2K, D, and L molecules (analogous to the human HLA-A, -B, and -C molecules), that is non-covalently assembled with the 12 kd monomorphic light chain, Beta2-microglobulin. Our objectives in these studies have been to examine a number of specific aspects of the biosynthesis, structure, and function of the class I molecule, including: 1) the measurement of the biochemical parameters that govern the binding of the MHC class I molecule to antigenic peptide; 2) the influence of Beta2-microglobulin and other serum factors on functional and biochemical binding of MHC class I molecules to antigenic peptides; 3) the expression, folding, assembly, structure, and stability of class I molecules produced in: a) an in vitro RNA-dependent translation system and b) an insect virus/moth ovary cell expression system; 4) the expression folding, assembly, structure, and stability of domain and sub-domain fragments of class I molecules generated by recombinant DNA methods, and expressed in tissue culture cells; 5) the function of soluble MHC class I molecules in a transgenic mouse model system; and 6) the production of single chain MHC class I molecules to examine the role of the heavy chain/Beta2-microglobulin interaction in the function and stability of these molecules and as the basis for high level production in prokaryotic systems.