ATTACHMENT OF RIBOSOMES TO MICROSOMAL MEMBRANES

该项目的长期目标是了解 亚细胞分布的生物遗传机制 膜和细胞器中的特定蛋白质。我们将专注于 在分子方面的机制决定了 关于膜的磷脂双层的配置 内质网合成的多肽。我们还将对排序进行研究 参与蛋白质在内质网中保留的过程 或确定将其转移到高尔基仪器，并从 在那里，到溶酶体或质膜。细胞色素P-450， 和内质网完整的膜蛋白，这是 共翻译插入膜中，但大部分仍保留 暴露在其细胞质表面，已被选为模式 其中一些工作所需的蛋白质。基本结构特征 使P-450中未切割的插入信号停止 远端序列的转移和将成熟蛋白锚定在 磷脂双层，将通过检查 嵌合多肽和修饰多肽的膜插入 在体外转录-翻译系统中产生 用基因工程模板编程。这些 研究还将提供对其他 未解锁的信号，不具有暂停传输功能，但 要么穿过薄膜，要么永久保留 固定在它上面。关于翻译后翻译的研究 对插入内质网的蛋白质进行分类，包括：1)分离 和移行池的生化特征 为蛋白质从内质网输送到 高尔基仪器。在这些研究中，合成一种 在这个细胞器中聚集的异常嵌合蛋白和 导致其过度开发将被采用。尝试次数将是 用来识别载体的受体，这些受体与 嵌合蛋白，可能会在这些过度发育的地方积累 水池。2)贝塔结构特征的识别 负责甘露糖磷酸化的葡萄糖醛酸苷酶 决定这种蛋白质在体内的滞留的残基 溶酶体。3)阐明途径，随后是 溶酶体膜蛋白与溶酶体膜蛋白的关系及鉴定 把这种蛋白质送到目的地的分拣信号。 4)确定导致保留的特征 细胞色素P-450在内质网细胞膜和微粒体内 内质网管腔内的酯酶。

The long term objective of this project is to understand the biogenetic mechanisms responsible for the subcellular distribution of specific proteins in membranes and organelles. We will focus on molecular aspects of the mechanisms that determine the disposition with respect to the phospholipid bilayer of membrane polypeptides synthesized in the ER. We will also study the sorting processes that are involved in the retention of proteins in the ER or determine their transfer to the Golgi apparatus, and from there, to lysosomes or the plasma membrane. Cytochrome P-450, and integral membrane protein of the ER which is cotranslationally inserted in the membrane but remains largely exposed on its cytoplasmic surface, has been chosen as a model protein for some of this work. The essential structural features that enable the uncleaved insertion signal in P-450 to halt transfer of distal sequences and to anchor the mature protein in the phospholipid bilayer, will be identified by examining the membrane insertion of chimeric and altered polypeptides generated in in vitro transcription-translation systems programmed with genetically engineered templates. These studies will also provide insights into the features of other uncleaved signals, which do not have halt transfer function but either pass through the membrane or remain permanently anchored to it. The studies concerning the posttranslational sorting of proteins inserted in the ER, include: 1) The isolation and biochemical characterization of the transitional cisternae that provide the site of exit for proteins delivered from the ER to the Golgi apparatus. for these studies, cells that synthesize an abnormal chimeric protein that accumulates in this organelle and leads to its overdevelopment will be employed. Attempts will be made to identify receptors of carriers that, complexed to the chimeric protein, may accumulate within these overdeveloped cisternae. 2) The identification of structural features in beta- glucuronidase responsible for the phosphorylation of the mannose residues that determine the sequestration of this protein in lysosomes. 3) an elucidation of the pathway followed by a lysosomal membrane protein to lysosomes and the identification of the sorting signal that address this protein to its destination. 4) An identification of the features that lead to the retention of cytochrome P-450 within the ER membrane and of a microsomal esterase within the ER lumen.