Biochemical characterization of a high affinity phosphate transporter (PiPT) from root endophyte fungus Piriformospora indica.

ABSTRACT

We have functionally characterized the high-affinity phosphate transporter (PiPT) from the root endophyte fungus Piriformospora indica. PiPT belongs to the major facilitator superfamily (MFS). PiPT protein was purified by affinity chromatography (Ni-NTA) and Size Exclusion Chromatography (SEC). The functionality of solubilized PiPT was determined in detergent-solubilized state by fluorescence quenching and in proteoliposomes. In the fluorescence quenching assay, PiPT exhibited a saturation concentration of approximately 2 µM, at a pH of 4.5. Proteoliposomes of size 121.6 nm radius, showed transportation of radioactive phosphate. Vmax was measured to be 232.2 ± 11 pmol/min/mg protein. We have found Km to be 45.8 ± 6.2 µM suggesting high affinity towards phosphate.