| The polyphosphate (PolyP) and phosphate (Pi) metabolisms of Deinococcus radiodurans R1 were characterized in view of engineering the radiation-resistant bacterium for radioactive metal biosequestration. Despite failure of the attempted approaches, fundamental insights were gained into the PolyP/Pi metabolisms, which may extend beyond Deinococcus. .;The D. radiodurans Pi-starvation inducible PHO regulon displayed a PhoB-mediated Gram-negative response, which demonstrated the singular ability to function in the absence of the high affinity Pst transporter and most likely independently of a PhoR sensor kinase. The PHO response was unaffected in a Deltappk mutant, but was constitutively low in Delta ppkDeltaphoU..;The PPK enzyme, which synthesizes PolyP reversibly from ATP, is responsible for all known high level PolyP accumulation in cells. PolyP synthesis is stimulated by high ATP levels, while the reverse activity is stimulated by high ADP levels. Unstressed D. radiodurans did not accumulate PolyP significantly, even upon ppk overexpression, which had proven successful in other systems, or upon deletion of genes encoding the main potential PolyP-utilizing enzymes As documented in other bacteria, the D. radiodurans Delta phoU displayed constitutively high PolyP levels and PHO induction. However, in the DeltappkDeltaphoU mutant, both were constitutively low.;The bacterium followed an overplus response, in which PolyP is accumulated after Pi starvation upon Pi re-addition. This PPK-dependent overplus response proved to create a key driving force for Pi uptake in Pi-starved cells. Pi-starved Delta pst took up Pi at a rate half that of the WT, while the Delta ppk mutant rate was ∼17 times lower. |
