# An Update on Polyphosphate In Vivo Activities
## 4 Bacteria  
In bacteria, polyP is synthesised and degraded by polyphosphate kinases (PPKs). Two distinct enzymes, PPK1 and PPK2 that differ in their reaction profile [41], reversibly catalyse the polymerisation of the terminal phosphate of ATP into a nascent polyP chain or  
\begin{table}
\begin{tabular}{l l l l l} \hline \hline
**Enzyme** & **Function** & **Regulation** & **Species** & **References** \\ \hline PPK1 & polyP polymerase using ATP & & Bacteria and _Dictyostelium_ & [39; 40] \\ \hline PPK2 & polyP polymerase using & & & \\  & GTP/ATP & & Bacteria in cytoplasm & [41] \\ \hline VTC & polyP polymerase using ATP & V-ATPase & & _S. cerevisiae_, Trypanosoma and Leishmana & [42; 43; 44] \\ \hline PPN1 & Endopolyphosphatase & & & _S. cerevisiae_, mammals & [45; 46] \\ \hline PPX1/2 & Exopolyn phosphatase & & Regulated by the accumulation of polyP in the cytosol & Fungi, _S. cerevisiae_ in mitochondria & [25; 47; 48; 49; 50] \\ \hline Nudt (Nudix hydrolase 15) & PolyP hydrolase & & Regulates oxidative stress in response to polyP & Mammals & [51] \\ \hline Prune/DRES17 & Short-chain & & Regulated by cytosolic & Mammalian and _Drosophila melanogaster_ cells in the cytosol & [49; 52; 53; 54] \\ \hline DDP1 (Diphosphinositol polyP Phosphohydrolase) & & & _S. cerevisiae_ in the cytosol & [45] \\ \hline Vtc4 (Vacuolar transporter chaperone-4) & polyP polymerase & not specified & _S. cerevisiae_ in Vacuole & [42; 55] \\ \hline PPN2 & & & Zn\({}^{2+}\) & _S. cerevisiae_ in acidocalcisomes & [56] \\ \hline PPX2 & Exopolyn phosphatase & Mg\({}^{2+}\), Mn\({}^{2+}\), and Ca\({}^{2+}\) & bacteria & [57] \\ \hline \hline \end{tabular}
\end{table}
Table 1: Major enzymes involved in metabolisms of polyP in different contemporary species.  
transfer terminal Pi from the polymer backbone to GDP, synthesising GTP [11; 58]. PPK2 is an 80 kDa enzyme bound to the inner leaflet of the plasma membrane and is evolutionarily older than PPK1 [59]. By modulating cellular energy status, polyP protects bacteria from various external stressors, including heat, ultraviolet irradiation, antibiotics, and oxidative stress. PPK1 deficiency by target ablation of ppk1 gene expression impairs growth and biofilm formation and reduces resistance to starvation [60; 61]. Moreover, gene ablation of _ppk2_ in _Mycobacterium smegmatis_ leads to biofilm synthesis and virulence [62; 63]. _Corynebacterium glutamic_ expressed two PPK2 isoenzymes, while _Pseudomonas aeruginosa_ and _Sinorthizobium melitoli_ have three isoenzymes each, and _Ralstonia eutropha_ even expresses five PPK2 variants [64]. Class I PPK2's phosphorylate nucleoside diphosphates ADP and GDP by transferring the terminal Pi of polyP to the nucleotide diphosphates and vice versa, shuttling Pi to the nascent polyP chain [65]. In contrast, class II PPK2's catalyse nucleotide monophosphate phosphorylation and were previously called polyP-AMP phosphotransferase (PAP) [66]. Class III PPK2's phosphorylate mono- or diphosphates in nucleotide to generate ADP or ATP, respectively, using polyP as a phosphate donor [67].  
Exopolynophatases (PPXs) hydrolyse polyP of chain length > 3 to release the terminal Pi. Based on the primary structure, they are classified into two types, PPX1 and PPX2, respectively. Both enzymes are members of the same protein family, the PPX-GppA phosphatases. Most bacteria, such as _Corynebacterium glutamicum_, express two distinct exopolynophatases. PPX1 and PPX2 from _C. glutamicum_ share low identity on the protein level (25%) and are different from enterobacterial, archaeal, and yeast exopolynophatases [57]. X-ray studies of the crystal structure of _E. coli_ PPX1 at 2.2-\({}^{\circ}\)A resolution revealed that the molecule is composed of four domains, with two of them mediating binding to polyP > 35-P\({}_{\text{i}}\) chain length and two harbouring the enzymatic activity, respectively. Bacterial PPX enzymes require both Mg\({}^{2+}\) and K\({}^{+}\) for maximal activity [68].