title: Structural analysis of the diadenylate cyclase reaction of DNA-integrity scanning protein A (DisA) and its inhibition by 3'-dATP.
abstract: The identification of the essential bacterial second messenger cyclic-di-AMP (c-di-AMP) synthesized by the DNA-integrity scanning protein A (DisA) has opened up a new and emerging field in bacterial signalling. To further analyse the diadenylate cyclase (DAC) reaction catalysed by the DAC domains of DisA, we crystallized Thermotoga maritima DisA in the presence of different ATP analogues and metal ions to identify the metal-binding site and trap the enzyme in pre- and post-reaction states. Through structural and biochemical assays we identified important residues essential for the reaction in the active site of the DAC domains. Our structures resolve the metal-binding site and thus explain the activation of ATP for the DAC reaction. Moreover, we were able to identify a potent inhibitor of the DAC domain. Based on the available structures and homology to annotated DAC domains we propose a common mechanism for c-di-AMP synthesis by DAC domains in c-di-AMP-producing species and a possible approach for its effective inhibition.
authors.affiliation: Gene Center and Department of Biochemistry, Ludwig-Maximilians-Universität, 81377 Munich, Germany.
authors.forename: Martina
authors.initials: M
authors.lastname: Müller
authors.affiliation: Gene Center and Department of Biochemistry, Ludwig-Maximilians-Universität, 81377 Munich, Germany.
authors.forename: Tobias
authors.initials: T
authors.lastname: Deimling
authors.affiliation: Gene Center and Department of Biochemistry, Ludwig-Maximilians-Universität, 81377 Munich, Germany Center for Integrated Protein Sciences, Ludwig-Maximilians-Universität, 81377 Munich, Germany.
authors.forename: Karl-Peter
authors.initials: KP
authors.lastname: Hopfner
authors.affiliation: Gene Center and Department of Biochemistry, Ludwig-Maximilians-Universität, 81377 Munich, Germany witte@genzentrum.lmu.de.
authors.forename: Gregor
authors.initials: G
authors.lastname: Witte
mesh_terms: D000255:Adenosine Triphosphate / Q000737:chemistry / Q000378:metabolism; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D001665:Binding Sites; D020134:Catalytic Domain; D000242:Cyclic AMP / Q000737:chemistry / Q000378:metabolism*; D008969:Molecular Sequence Data; D019761:Phosphorus-Oxygen Lyases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D016415:Sequence Alignment; D020124:Thermotoga maritima / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics
chemical_list: D001426:Bacterial Proteins; D000255:Adenosine Triphosphate; D000242:Cyclic AMP; D019761:Phosphorus-Oxygen Lyases
keywords: crystal structure; cyclic-di-adenosine 5′-monophos-phate (c-di-AMP); diadenylate cyclase (DAC) domain; inhibition; reaction mechanism