Inorganic Polyphosphate, Exopolyphosphatase, and Pho84 -Like Transporters May Be Involved in Copper Resistance in Metallosphaera sedula DSM 5348 T .

Polyphosphates (PolyP) are linear polymers of orthophosphate residues that have been proposed to participate in metal resistance in bacteria and archaea. In addition of having a CopA/CopB copper efflux system, the thermoacidophilic archaeon Metallosphaera sedula contains electron-dense PolyP-like granules and a putative exopolyphosphatase (PPX Msed , Msed_0891 ) and four presumed pho84 -like phosphate transporters ( Msed_0846 , Msed_0866 , Msed_1094 , and Msed_1512 ) encoded in its genome. In the present report, the existence of a possible PolyP-based copper-resistance mechanism in M. sedula DSM 5348T was evaluated. M. sedula DSM 5348T accumulated high levels of phosphorous in the form of granules, and its growth was affected in the presence of 16 mM copper. PolyP levels were highly reduced after the archaeon was subjected to an 8 mM CuSO4 shift. PPX Msed was purified, and the enzyme was found to hydrolyze PolyP in vitro . Essential residues for catalysis of PPX Msed were E111 and E113 as shown by a site-directed mutagenesis of the implied residues. Furthermore, M. sedula ppx , pho84 -like, and copTMA genes were upregulated upon copper exposure, as determined by qRT-PCR analysis. The results obtained support the existence of a PolyP-dependent copper-resistance system that may be of great importance in the adaptation of this thermoacidophilic archaeon to its harsh environment.