LI FANGYANBIOLOGICAL SCIENCES2010-04-082010-04-082007-04-26LI FANGYAN (2007-04-26). The role of a novel second messenger, c-di-GMP in pseudomonas putida via enzymatic study of MorA. ScholarBank@NUS Repository.https://scholarbank.nus.edu.sg/handle/10635/16190MorA is a membrane-localized regulator that contains GGDEF and EAL domains implicated in controlling c-di-GMP levels via diguanylate cyclase (DGC) and phosphodiesterase (PDE) activities, respectively. MorA controls the timing of flagellar development and affects motility, chemotaxis, and biofilm formation in Pseudomonas species. Due to the presence of GGDEF and EAL domains, we proposed that MorA acts via modulating intracellular c-di-GMP levels. A method for extraction and detection of c-di-GMP was established in Gluconacetobacter and Pseudomonas. Comparison of c-di-GMP levels in P. putida wild type (WT), morA knock out (KO) and overexpressed (OE) strains showed that MorA negatively affects c-di-GMP levels. Enzymatic activity assay suggested MorA does not possess DGC activity. Hence, it is likely to affect via PDE activity. With the purpose of investigating factors affecting c-di-GMP metabolism by availability of free GTP as substrate of cyclase activity, four putative GTP-binding proteins were identified and characterized in P. putida.enc-di-GMP, diguanylate cyclase, phosphodiesterase, GGDEF, EAL, MorAThesisNOT_IN_WOS