Effect of speciation of uranium on localisation of uranyl phosphate precipitate in recombinant D. radiodurans expressing PhoN/PhoK

Phosphatase mediated enzymatic bioprecipitation of uranium (U) as insoluble uranyl phosphate is an attractive bioremediation strategy. Earlier PhoN (a non specific acid phosphatase) was cloned and over-expressed in Deinococcus radiodurans to achieve bioprecipitation of U from acidic to neutral conditions. Similarly, a PhoK gene encoding an alkaline phosphatase has now been cloned and overexpressed in this microbe for bioprecipitation of U from alkaline waste solutions. In case of Deino-PhoK strain, a high phosphatase activity was detected in the spent medium, while Deino-PhoN cells did not show any activity in the spent medium, suggesting that PhoK is extracellular in nature while PhoN is not. Deino-PhoN cells, when incubated with 1 mM uranyl nitrate (pH 6.8) and 5 mM β-GP as substrate, showed cell surface bound spicule-like precipitate in SEM/TEM. In contrast Deino-PhoK cells, exposed to 1 mM uranyl carbonate (pH 9) and 5 mM β-GP, showed extracellular precipitate in solution. Localisation of uranyl phosphate precipitate depended on the precipitation conditions and Deino-PhoN cells displayed distinct extracellular precipitate crystals with uranyl carbonate (pH 9), while Deino-PhoK cells clearly displayed cell surface bound crystals of uranyl phosphate, when employed with uranyl nitrate (pH 6.8). These results suggest that differential localisation of precipitate may not be affected by localisation of the enzyme alone, but is mainly governed by the species of uranium being used (which is also dependent on pH). This work provides a graphic understanding of pH-dependent interactions of uranium with D. radiodurans cell surfaces. (author)