title: The PilZ domain is a receptor for the second messenger c-di-GMP: the PilZ domain protein YcgR controls motility in enterobacteria.
abstract: The ubiquitous bacterial second messenger c-di-GMP controls exopolysaccharide synthesis, flagella- and pili-based motility, gene expression, and interactions of bacteria with eukaryotic hosts. With the exception of bacterial cellulose synthases, the identities of c-di-GMP receptors and end targets have remained unknown. Recently, Amikam and Galperin (Amikam, D., and Galperin, M. (2006) Bioinformatics 22, 3-6) hypothesized that the PilZ domains present in the BcsA subunits of bacterial cellulose synthases function in c-di-GMP binding. This hypothesis has been tested here using the Escherichia coli PilZ domain protein YcgR, its individual PilZ domain and the PilZ domain from Gluconacetobacter xylinus BcsA. YcgR was purified and found to bind c-di-GMP tightly and specifically, Kd 0.84 microm. Individual PilZ domains from YcgR and BcsA also bound c-di-GMP, albeit with lesser affinity, indicating that PilZ is sufficient for binding. The site-directed mutagenesis performed on YcgR implicated the most conserved residues in the PilZ domain directly in c-di-GMP binding. It is suggested that c-di-GMP binding to PilZ brings about conformational changes in the protein that stabilize the bound ligand and initiate the downstream signal transduction cascade. While the identity of the downstream partner(s) of YcgR remains unknown, it is shown that YcgR regulates flagellum-based motility in a c-di-GMP-dependent manner. The inactivation of ycgR improves swimming and swarming motility of the poorly motile yhjH mutants of Salmonella enterica serovar Typhimurium UMR1. Therefore, biochemical and genetic evidence presented here establishes PilZ as a long sought after c-di-GMP-binding domain and YcgR as a c-di-GMP receptor affecting motility in enterobacteria.
authors.affiliation: Department of Molecular Biology, University of Wyoming, Laramie, Wyoming 82071, USA.
authors.forename: Dmitri A
authors.initials: DA
authors.lastname: Ryjenkov
authors.forename: Roger
authors.initials: R
authors.lastname: Simm
authors.forename: Ute
authors.initials: U
authors.lastname: Römling
authors.forename: Mark
authors.initials: M
authors.lastname: Gomelsky
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry; D002352:Carrier Proteins / Q000737:chemistry; D006152:Cyclic GMP / Q000031:analogs & derivatives* / Q000737:chemistry; D004755:Enterobacteriaceae / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D041822:Gluconacetobacter xylinus / Q000378:metabolism; D007700:Kinetics; D008958:Models, Molecular; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary
chemical_list: D001426:Bacterial Proteins; D002352:Carrier Proteins; D029968:Escherichia coli Proteins; C415947:YcgR protein, E coli; C062025:bis(3',5')-cyclic diguanylic acid; D006152:Cyclic GMP