Generation and application of second messenger molecules by SMODS and SAVED

项目总结/摘要 基于核苷酸的第二信使在生物体中发挥重要的生物学功能。之间 各种各样的第二信使分子，其中几种是由属于 Polb超家族在动物中，胞质NTases OAS和cGAS产生2 '-5'-连接的寡腺苷酸和2 '-连接的寡腺苷酸。 5 '-连接的c-GAMP，两者都能触发针对病毒和细菌的先天免疫应答 感染在细菌中，NTase DncV产生3 '-5'-连接的cGAMP，其激活CapV以抑制细胞增殖。 增长利用比较基因组学、序列保守性和结构分析的组合， Aravind及其同事发现了细菌中以核苷酸为中心的系统的巨大网络。口之家 被称为SMODS（第二信使寡核苷酸和二核苷酸合成酶）的NTases被预测为 生成第二信使。并预测了几个保守结构域可能是该蛋白的受体。 第二信使由SMODS产生。在预测的受体中，名为SAVED的结构域 （SMODS相关的和融合到各种效应结构域）是最丰富的。除了一个 最近的表征所产生的产品之一的SMODS，但是，预测的生化 SMODS和SAVED的生物学功能尚未得到实验验证。采用 生物信息学，生物化学和结构生物学的方法，我们的目标是追求以下两条路线 SMODS和SAVED的研究：1）我们将在体外重建SMODS的酶活性， 探测SAVED和SMODS产生的第二信使之间的相互作用; 2）我们将携带 SMODS和SAVED的结构研究，SMODS与其活化剂和底物的复合物，以及 与SMODS产生的第二信使复合保存。

Project Summary/Abstract Nucleotide-based second messengers play important biological functions in living organisms. Among various second messenger molecules, several of them are generated by families of NTases that belong to Polb superfamily. In animals, cytosolic NTases OAS and cGAS generate 2’-5’-linked oligoadenylates and 2’- 5’-linked c-GAMP, respectively, both of which trigger innate immune response against viral and bacterial infection. In bacteria, NTase DncV generates 3’-5’-linked cGAMP, which activates CapV to inhibit cell growth. Using a combination of comparative genomics, sequence conservation, and structure analysis, Aravind and coworkers uncovered a vast network of nucleotide-centric systems in bacteria. A family of NTases named SMODS (secondary messenger oligo and dinucleotides synthase) was predicted to generate second messengers. And several conserved domains were predicted to be receptors of the second messengers generated by SMODS. Among the predicted receptors, the domain named SAVED (SMODS-associated and fused to various effector domains) is the most abundant. With the exception of a recent characterization of the product generated by one of SMODS, however, the predicted biochemical and biological functions of SMODS and SAVED have not been experimentally tested. Employing approaches of bioinformatics, biochemistry, and structural biology, we aim to pursue the following two lines of investigation of SMODS and SAVED: 1) We will in vitro reconstitute the enzymatic activity of SMODS and probe interaction between SAVED and the second messengers generated by SMODS; and 2) We will carry out structural studies of SMODS and SAVED, SMODS in complex with its activator and substrates, and SAVED in complex with the second messengers generated by SMODS.