IMPORT OF POLYPEPTIDES INTO CHLOROPLASTS

核酮糖-1，5-二磷酸羧化酶的小亚基（SS）和 捕光叶绿素a/B的组分（AB）多肽 蛋白质复合物作为前体在细胞质核糖体上合成 多肽。 这些前体（preSS和preAB）的特征在于 N-末端延伸称为转运肽。 我们最近 证明前SS转运肽可以介导 外源多肽翻译后导入叶绿体。 这 一项建议涉及到界定这一观察的普遍性 以及转运肽内的序列，其介导个体 进口的步骤。 对于这些研究，我们将使用体外表达系统 以产生标记的目的多肽。 多肽，我们 建议研究包括多种融合多肽的区分 特征和突变多肽， preSS或preAB的转运肽内的缺失。 我们将 检查单个多肽输入分离的叶绿体 对于那些有缺陷的，我们将确定缺陷是否 发生在绑定、移位或处理中。 通过这些程序，我们 期望表征转运肽内的结构域， 目的地导入过程。 我们将在这些域中定义序列 通过阿托伐他汀定向诱变。 拟议的研究将 有助于我们了解多肽的输入过程 细胞器，并将有助于未来的操纵表型 植物的特征。

The small subunit (SS) of ribulose-1,5-bisphosphate carboxylase and the constituent (AB) polypeptides of the light-harvesting chlorophyll a/b protein complex are synthesized on cytoplasmic ribosomes as precursor polypeptides. These precursors (preSS and preAB) are characterized by an N-terminal extension known as the transit peptide. We have recently demonstrated that the transit peptide for preSS can mediate the posttranslational import into chloroplasts of a foreign polypeptide. This proposal is concerned with defining both how general this observation is and the sequences within the transit peptide that mediate the individual steps of import. For these studies we will use in vitro expression systems to generate the labeled polypeptides of interest. The polypeptides we propose to study include a variety of fusion polypeptides of distinguishing characteristics and mutant polypeptides in which we will have generated deletions within the transit peptide of either preSS or preAB. We will examine the import into isolated chloroplasts for individual polypeptides and for those that are defective we will determine whether the defect occurs with bindings, translocation or processing. By these procedures we expect to characterize domains within the transit peptides that mediate destinct import processes. We will define sequences within these domains by oligonucleotide-directed mutagenesis. The proposed studies will contribute to our understanding of the process of import of polypeptides into organelles and will aid the future manipulation of phenotypic characteristics of plants.