Structural biochemistry of c-di-AMP degrading phosphodiesterases and their regulation

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Nucleotide second messengers are key components of signal transduction events that link external/internal signals to the cellular response of the cell. The discovery of c-di-AMP as a new and also essential bacterial second messenger opened up a new field in bacterial signaling. Whereas a set of proteins in the c-di-AMP pathway has been already been characterized, the interplay of proteins and connectivity in the c-di-AMP regulatory processes is yet still unclear. Due to the fact that c-di-AMP is essential (for the bacteria synthesizing it) and the fact that changes in the c-di-AMP level have dramatic effect on the cells, altering the concentration of c-di-AMP might be a potential target in antimicrobial therapy. We aim to analyze the degradation of c-di-AMP by specific phosphodiesterases (PDE) carrying a DHH-DHHA1-domain that is responsible for the hydrolysis of the cyclic dinucleotide. Interestingly, two subfamilies of these DHH-type PDE have been described, one membrane protein-type (GdpP) possessing also additional regulatory domains (GGDEF and PAS) and one cytosolic PDE only comprising the DHH-DHHA1-domain. Whereas the first group degrades c-di-AMP to 5-pApA, the latter is able to degrade both c-di-AMP but also 5-pApA to AMP. We aim to use biochemical, biophysical and structural methods (supported by in vivo methods) to elucidate the structure function relationship of these two types of PDEs. We will try to identify the structural differences between the two DHH-type PDEs leading to different products and also try to shed light on the regulation of GdpP-type PDEs by its accessory domains (GGDEF and PAS).