Characterization and analysis of a novel diguanylate cyclase PA0847 from Pseudomonas aeruginosa PAO1

Yan Zhang,1,2 Jiayi Guo,1 Ning Zhang,2 Wensu Yuan,2 Zhi Lin,2 Weidong Huang11Department of Biochemistry and Molecular Biology, School of Basic Medicine, Ningxia Medical University, Yinchuan 750004, People’s Republic of China; 2School of Life Sciences, Tianjin University, Tianjin 300072, People’s Republic of ChinaBackground: As a central signaling molecule, cyclic diguanylate (c-di-GMP) is found to regulate various bacterial phenotypes, especially those involved in pathogen infection and drug resistance. Noticeably, many microbes have up to dozens of proteins that are involved in c-di-GMP metabolism. This apparent redundancy and the relevant functional specificity have become the focus of research. While a number of these proteins have been identified and investigated, the functions of PA0847, a PAS and GGDEF domain-containing protein from Pseudomonas aeruginosa PAO1, remain unclear.Materials and methods: In the current study, microbiology, biochemistry and structural biology methods were applied to characterize the gene/protein of PA0847.Results: We showed that PA0847 affects bacterial motility but not biofilm formation. We recorded the phenotypic influences of amino acids and compounds, and found that PA0847 is involved in response to various environmental nutrients and factors, suggesting its possible role in sensing environmental cues. Both in-vitro and in-vivo studies showed that PA0847 is an active diguanylate cyclase (DGC), whose activity depends on the neighboring PAS domain. Interestingly, PA0847 demonstrates no significant product inhibition, though the key residues of two I-sites for c-di-GMP binding are conserved in its GGDEF domain. A local structural change imposed by an adjacent tyrosine residue was identified, which indicates the structural and functional diversities of the GGDEF family proteins.Conclusion: Our data provide evidence for understanding the signaling mechanism of the unique c-di-GMP metabolizing protein PA0847.Keywords: Pseudomonas aeruginosa, c-di-GMP, GGDEF domain, diguanylate cyclase, structur