## 11 PgpH and GdpP-Type Phosphodiesterases  
PgpH-type phosphodiesterase were first identified in _Listeria monocytogenes_ and are widespread among Firmicutes, though absent in _Staphylococcus_ and _Streptococcus_ species [81; 84; 105]. PgpH contains eight transmembrane helices, an extracellular seven transmembrane helix-HDED domain (7 TMR-HDED) domain, seven transmembrane helices, and a cytosolic HD phosphohydrolase domain, which hydrolyzes c-di-AMP into pApA (Figure 8A) [94]. Crystal Structure of _L monocytogenes_ PDE PgpH HD domain in complex with Cyclic-di-AMP give insight into the catalytic mechanisms of PgpH PDE (Figure 10). Structural analysis reveals that in the Fe-bound active site, a bridging water molecule is positioned beneath the 5\({}^{\prime}\) phosphate of nucleotide, the target of hydrolysis. This water is activated by the two metal ions and a hydrogen bond with Asp648, which also coordinates Fe1. The water/hydroxide then acts as a nucleophile, attacking the phosphorus atom, resulting in the release of the 3\({}^{\prime}\) hydroxyl of nucleotide 2. The oxyanion formed is stabilized by the ammonium group of Lys547, which, along with a solvent water molecule, facilitates protonation to complete the reaction (Figure 10B). Given the conserved nature  
Figure 9: (**A**) Structures of small molecules inhibitors of NmA discovered by the Sintim group (**C82**) [97], the Bishai group (**C-33** and **C-34**) [99], and Stingray Therapeutics (**I-7**) [98]. (**B**) The DhhP knockout strain of Borrelia burgdorferi shows growth defects; imaged with darkfield microscopy. (**C**) Quantification of cell length from 100 randomly selected cells from (**B**). Reproduced with permission from [83]. Copyright © 2014 American Society for Microbiology-Journals.  
of the HD domain, this catalytic mechanism is likely shared among other HD-domain phosphodiesterase.  
The GdpP-type phosphodiesterase was first identified in _Lactococcus lactis_ and later found in various Firmicutes, including _B. subtilis_, _S. aureus_, _S. pneumoniae_, and _Enterococcus_ species [77, 79, 82, 83, 85, 106, 107, 85, 108, 109]. GdpP contains transmembrane helices, a PAS domain, a degenerated GGDEF domain and a DHH/DHHA1 phosphodiesterase domain, which degrades c-di-AMP into pApA (Figure 8A) [105, 110]. The catalytic mechanism of GdpP PDEs are the same with DhhP since they both contain DHH/DHH1 domains [111]. GdpP is regulated by heme and nitric oxide (NO), which inhibit its activity, as well as the bacterial stress signal (p)ppGpp, which competitively inhibits its catalytic function [109, 112]. In many Firmicutes, GdpP inactivation leads to elevated c-di-AMP levels, resulting in cell wall-damaging antibiotics resistance and altered virulence [81, 112]. However, studies by [PERSON] et al. have demonstrated that a _gdpP pgpH_ double mutant is toxic to _Bacillus subtilis_, prompting the inactivation of diadenylate cyclase AdaA to counterbalance c-di-AMP accumulation [81]. Similarly, research by [PERSON] et al. [94] found that a _gdpP pgpH_ double mutant in _Listeria monocytogenes_ exhibited enhanced host inflammation, intracellular growth defects, and significantly reduced virulence in a murine infection model, suggesting that dual inhibitors of GdpP and PgpH could serve as anti-virulence agents against Listeria infection.