%0 Journal Article %1 Düvel31082015 %A Düvel, J %A Bense, S %A Möller, S %A Bertinetti, D %A Schwede, F %A Morr, M %A Eckweiler, D %A Genieser, H G %A Jänsch, Lothar %A Herberg, Friedrich W. %A Frank, Ronald %A Häussler, Susanne %D 2015 %E Bacteriol, J %J Journal of Bacteriology %K haeussler %N 1 %P 138-146 %T Application of synthetic peptide arrays to uncover c-di-GMP binding motifs %V 198 %X High levels of the universal bacterial second messenger c-di-GMP promote the establishment of surface–attached growth in many bacteria. C-di-GMP can not only bind to nucleic acids and directly control gene expression, but it also binds to a diverse array of proteins of specialized functions and orchestrates their activity. Since its development in the early 90s the synthetic peptide array technique has become a powerful tool for high throughput approaches and was successfully applied to investigate the binding specificity of protein-ligand interactions. In this study, we used peptide arrays to uncover the c-di-GMP binding site of a Pseudomonas aeruginosa protein (PA3740) that was isolated in a chemical proteomics approach. PA3740 was shown to bind c-di-GMP with high affinity and peptide arrays uncovered LKKALKKQTNLR as a putative c-di-GMP binding motif. Most interestingly, different from the previously identified c-di-GMP binding motif of the PilZ domain (RxxxR) or the I-site of diguanylate cyclases (RxxD), not the charged amino acids provided the key residues of the binding sequence but two leucine residues and a glutamine residue. Those three amino acids are highly conserved across PA3740 homologs and their singular exchange to alanine reduced c-di-GMP binding within the full length protein.Importance In many bacterial pathogens the universal bacterial second messenger c-di-GMP governs the switch from the planktonic, motile to the sessile, biofilm mode of growth. Bacteria adapt their intracellular c-di-GMP levels to a variety of environmental challenges. Several classes of c-di-GMP binding proteins have been structurally characterized, and diverse c-di-GMP binding domains were identified. Nevertheless for several c-di-GMP receptors the binding motif remains to be determined. Here we show that the use of a synthetic peptide array allowed the identification of a c-di-GMP binding motif of a putative c-di-GMP receptor protein in the opportunistic pathogen P. aeruginosa. The application of synthetic peptide arrays will facilitate the search for additional c-di-GMP receptor proteins and aid in the characterization of c-di-GMP binding motifs.

@article{Düvel31082015, abstract = {High levels of the universal bacterial second messenger c-di-GMP promote the establishment of surface–attached growth in many bacteria. C-di-GMP can not only bind to nucleic acids and directly control gene expression, but it also binds to a diverse array of proteins of specialized functions and orchestrates their activity. Since its development in the early 90s the synthetic peptide array technique has become a powerful tool for high throughput approaches and was successfully applied to investigate the binding specificity of protein-ligand interactions. In this study, we used peptide arrays to uncover the c-di-GMP binding site of a Pseudomonas aeruginosa protein (PA3740) that was isolated in a chemical proteomics approach. PA3740 was shown to bind c-di-GMP with high affinity and peptide arrays uncovered LKKALKKQTNLR as a putative c-di-GMP binding motif. Most interestingly, different from the previously identified c-di-GMP binding motif of the PilZ domain (RxxxR) or the I-site of diguanylate cyclases (RxxD), not the charged amino acids provided the key residues of the binding sequence but two leucine residues and a glutamine residue. Those three amino acids are highly conserved across PA3740 homologs and their singular exchange to alanine reduced c-di-GMP binding within the full length protein.Importance In many bacterial pathogens the universal bacterial second messenger c-di-GMP governs the switch from the planktonic, motile to the sessile, biofilm mode of growth. Bacteria adapt their intracellular c-di-GMP levels to a variety of environmental challenges. Several classes of c-di-GMP binding proteins have been structurally characterized, and diverse c-di-GMP binding domains were identified. Nevertheless for several c-di-GMP receptors the binding motif remains to be determined. Here we show that the use of a synthetic peptide array allowed the identification of a c-di-GMP binding motif of a putative c-di-GMP receptor protein in the opportunistic pathogen P. aeruginosa. The application of synthetic peptide arrays will facilitate the search for additional c-di-GMP receptor proteins and aid in the characterization of c-di-GMP binding motifs.}, added-at = {2015-09-09T11:57:41.000+0200}, author = {Düvel, J and Bense, S and Möller, S and Bertinetti, D and Schwede, F and Morr, M and Eckweiler, D and Genieser, H G and Jänsch, Lothar and Herberg, Friedrich W. and Frank, Ronald and Häussler, Susanne}, biburl = {https://www.bibsonomy.org/bibtex/2ffaf7cc7d035424f8329ebe23e79d8d3/haeussler}, editor = {Bacteriol, J}, interhash = {45a33b95818a60e156c615b66551fa35}, intrahash = {ffaf7cc7d035424f8329ebe23e79d8d3}, journal = {Journal of Bacteriology}, keywords = {haeussler}, number = 1, pages = {138-146}, pubmedurl = {http://www.ncbi.nlm.nih.gov/pubmed/26324453}, timestamp = {2016-01-11T12:05:43.000+0100}, title = {Application of synthetic peptide arrays to uncover c-di-GMP binding motifs}, volume = 198, year = 2015 }