Structural Basis of Activity and Allosteric Control of Diguanylate Cyclase

• Published in: Proceedings of the National Academy of Sciences - PNAS 2004-12, Vol.101 (49), p.17084-17089
• Main Authors: Chan, Carmen, Paul, Ralf, Samoray, Dietrich, Amiot, Nicolas C., Giese, Bernd, Jenal, Urs, Schirmer, Tilman, Deisenhofer, Johann
• Format: Article
• Language: English
• Subjects: Active sites; Allosteric Regulation; Bacterial Proteins - chemistry; Bacterial Proteins - metabolism; Binding Sites; Biological Sciences; Biophysics; Catalysis; Caulobacter crescentus; Caulobacter crescentus - enzymology; Crystal structure; Crystallography; Crystallography, X-Ray; Crystals; Dimers; Enzymes; Escherichia coli Proteins; Ligands; Models, Molecular; Molecular Structure; Molecules; Monomers; Phosphorus-Oxygen Lyases - chemistry; Phosphorus-Oxygen Lyases - metabolism; Phosphorylation; Protein Binding; Protein Conformation
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.0406134101

Recent discoveries suggest that a novel second messenger, bis-(3′→5′)-cyclic di-GMP (c-diGMP), is extensively used by bacteria to control multicellular behavior. Condensation of two GTP to the dinucleotide is catalyzed by the widely distributed diguanylate cyclase (DGC or GGDEF) domain that occurs in various combinations with sensory and/or regulatory modules. The crystal structure of the unorthodox response regulator PleD from Caulobacter crescentus, which consists of two CheY-like receiver domains and a DGC domain, has been solved in complex with the product c-diGMP. PleD forms a dimer with the CheY-like domains (the stem) mediating weak monomer-monomer interactions. The fold of the DGC domain is similar to adenylate cyclase, but the nucleotide-binding mode is substantially different. The guanine base is H-bonded to Asn-335 and Asp-344, whereas the ribosyl and α-phosphate moieties extend over the β2-β3-hairpin that carries the GGEEF signature motif. In the crystal, c-diGMP molecules are crosslinking active sites of adjacent dimers. It is inferred that, in solution, the two DGC domains of a dimer align in a two-fold symmetric way to catalyze c-diGMP synthesis. Two mutually intercalated c-diGMP molecules are found tightly bound at the stem-DGC interface. This allosteric site explains the observed noncompetitive product inhibition. We propose that product inhibition is due to domain immobilization and sets an upper limit for the concentration of this second messenger in the cell.