STRUCTURAL BASIS OF RNA-CATALYZED AMINOACYLATION OF RNA

描述（由申请人提供）：氨基酸与接头RNA分子的连接是蛋白质生物合成中的关键反应。氨酰化反应在细胞中由氨酰-tRNA合成酶催化。最近，已经通过体外选择获得了三类催化RNA（核酶），其通过将氨酰基部分从预先形成的腺苷酸转移到其自身的3 '-末端来重现这种活性。第一类包含接受任何氨酰基部分的RNA，而第二类的成员对苯丙氨酸和酪氨酸具有特异性。第三类核酶也催化二肽的形成，这一反应类似于由大型核酶（核糖体）催化的反应。在这个提议中，这三类自氨酰化RNA的三维结构将通过X射线晶体学来确定。核酶将与底物和产物类似物复合分析。本研究中获得的结构信息将揭示这些核酶的活性位点的构型、氨基酸特异性结合口袋的组织和催化机制。这将有助于澄清氨基酰基转移到肽或RNA是否涉及熵催化（即，它需要酶上的特殊催化基团，如酸或碱;因此，它将增加我们对较大生物系统（如氨酰-tRNA合成酶和核糖体）催化反应的理解。

DESCRIPTION (provided by applicant): Attachment of amino acids to adapter RNA molecules is a key reaction in protein biosynthesis. The aminoacylation reaction is catalyzed in the cell by aminoacyl-tRNA synthetases. Recently, three classes of catalytic RNAs (ribozymes) that recapitulate this activity by transferring the aminoacyl moiety from a pre-formed adenylate to their own 3'-termini have been obtained through in vitro selection. The first class contains RNAs that accept any aminoacyl moiety, whereas members of the second class are specific for phenylalanine and tyrosine. Ribozymes of the third class also catalyze the formation of dipeptides, a reaction analogous to that catalyzed by a large ribozyme, the ribosome. In this proposal, the three-dimensional structures of self-aminoacylating RNAs of the three classes will be determined by x-ray crystallography. The ribozymes will be analyzed in complex with substrate and product analogs. The structural information obtained in this study will reveal the configuration of the active site of these ribozymes, organization of the amino acid-specific binding pocket and the mechanism of catalysis. It will help clarify the issue of whether the aminoacyl transfer to a peptide or an RNA involves entropic catalysis (i.e., precise positioning of the substrates) or it requires special catalytic groups on the enzyme, such as an acid or base; hence, it will add to our understanding of the reactions catalyzed by the larger biological systems such as the aminoacyl-tRNA synthetases and the ribosome.