铜绿假单胞菌PAO1中环鸟苷二磷酸代谢蛋白PA2072的生物学功能及信号转导作用的研究

近年来，人们发现，新型小分子环核苷酸第二信使—环鸟苷二磷酸(3',5'-cyclic-di-guanosine monophosphate, c-di-GMP)能够在转录、转录后及翻译后水平上，对细菌的生物膜合成、运动性、耐药性、致病性，细胞周期及分化等一系列生理过程，起到重要的调控作用。在细菌胞内，环鸟苷二磷酸由含有GGDEF结构域的鸟苷酸环化酶催化合成，而由含有EAL结构域的酯酶催化分解。预实验结果提示，铜绿假单胞菌PAO1中未经报道的GGDEF-EAL结构域基因PA2072具有复杂的生物学功能。在本项申请中，我们将对PA2072展开较为系统深入的研究：利用生物化学、分子生物学、结构生物学等实验方法，明确PA2072的催化功能；分析其对环境信号的感应；研究其与其它环鸟苷二磷酸代谢蛋白的相互作用。我们的工作将深化对于环鸟苷二磷酸信号转导作用机制的认识，为未来控制重要致病菌的感染奠定基础。

Cyclic dimeric guanosine monophosphate (c-di-GMP), a universally conserved second messenger, has been studied very actively in recent years. C-di-GMP is involved in regulation of various bacteria physiological functions, including the bacterial motility and biofilm formation. Most noticeably, it contributes to the pathogenicity of numerous bacterial species..In bacteria cells, c-di-GMP is synthesized from two molecules of GTP by diguanylate cyclases (DGCs, GGDEF domain containing proteins), and degraded to pGpG by phosphodiesterases (PDEs, EAL domain containing proteins). These enzymes are therefore designated as c-di-GMP metabolizing proteins. .Pseudomonas aeruginosa, a widespread Gram-negative bacterium, is an important model organism and can cause a variety of acute and chronic infections. .In current proposal, we will focus on the uncharacterized c-di-GMP metabolizing protein of PA2072 from Pseudomonas aeruginosa PAO1. Mainly through biochemical, molecular biology and structural biology techniques, we will analyze the C-terminal catalytic activity, and identify the N-terminal sensor molecules of PA2072. We will also study the protein-protein interactions of PA2072 with other c-di-GMP metabolizing proteins. Our work will not only dissect the biological functions of the gene of PA2072, but also help to deepen our understandings of the mechanisms of c-di-GMP signaling.