CHARACTERIZATION OF MAMMALIAN MITOCHONDRIAL RIBOSOMES

哺乳动物线粒体含有55 S核糖体的不寻常的组成， 物理化学性质 拟议研究的目标是 的结构、组成、功能和生物合成的阐明 哺乳动物线粒体核糖体。 从牛肝中大规模生产的线粒体核糖体正在被 其特征在于结构和功能，以提供用于 哺乳动物线粒体核糖体的研究。 单个的r-蛋白将 根据它们的分子量、等电点、化学计量、氨基 酸组成，N-末端氨基酸，胰蛋白酶肽指纹，和 N-末端氨基酸序列。 审查结构的方法 线粒体R蛋白的组织将包括： 单核糖体和亚核糖体颗粒中的r-蛋白与表面探针的相互作用， 鉴定“核心”颗粒中的r-蛋白和 亚核糖体颗粒 识别单个R蛋白的方法 定义的功能域将包括蛋白质的功能重建 缺陷的亚核糖体颗粒和位点特异性亲和标记 probes. 哺乳动物线粒体r蛋白的种间变异将 来评估它们的快速进化分化。 实验将 也可以进行以确定哪些线粒体R蛋白是在 线粒体，并表征细胞质核糖体参与 线粒体蛋白质的合成。

Mammalian mitochondria contain 55S ribosomes of unusual composition and physical-chemical properties. The goal of the proposed research is the elucidation of the structure, composition, function, and biosynthesis of mammalian mitochondrial ribosomes. Mitochondrial ribosomes produced on a large scale from bovine liver are being characterized structurally and functionally to provide a model system for the study of mammalian mitochondrial ribosomes. The individual r-proteins will be characterized with respect to their MW, isoelectric point, stoichiometry, amino acid composition, N-terminal amino acid, tryptic peptide fingerprint, and N-terminal amino acid sequence. Approaches to examine the structural organization of mitochondrial r-proteins will include: the accessibility of r-proteins in monoribosomes and subribosomal particles to surface probes and the identification of r-proteins in "core" particles and fragments of the subribosomal particles. Approaches to identify individual r-proteins within defined functional domains will include the functional reconstitution of protein deficient subribosomal particles and labelling with site-specific affinity probes. The interspecific variation of mammalian mitochondrial r-proteins will be analyzed to assess their rapid evolutionary divergence. Experiments will also be performed to ascertain which of the mitochondrial r-proteins are made in mitochondria and to characterize the cytoplasmic ribosomes involved in the synthesis of mitoribosomal proteins.