Cell surface-expression of the phosphate-binding protein PstS: System development, characterization, and evaluation...

Simultaneous overabundance and scarcity of inorganic phosphate (P_i) is a critical issue driving the development of innovative water/wastewater treatment technologies that not only facilitate P_i removal to prevent eutrophication, but also recover P_i for agricultural reuse. Here, a cell-surface expressed high-affinity phosphate binding protein (PstS) system was developed, and its P_i capture and release potential was evaluated. E. coli was genetically modified to express PstS on its outer membrane using the ice nucleation protein (INP) as an anchoring motif. Verification of protein expression and localization were performed utilizing SDS-polyacrylamide gel electrophoresis (SDS-PAGE), western blot, and outer membrane separation analyses. Cell surface characterization was investigated through acid-base titration, X-ray photoelectron spectroscopy (XPS), and Fourier transform infrared spectroscopy (FTIR). These tests provided information on the macromolecular structure and composition of the bacteria surface as well as the proton-exchange properties of the surface functional groups (i.e., pKa values). Phosphate desorption and adsorption batch experiments were conducted to evaluate the effects of temperature, pH, and ionic strength on phosphate capture and release. The PstS surface-displayed cells demonstrated greater potential to release and capture phosphate compared to non-modified cells. Higher temperatures up to 40°C, basic pH conditions (pH = 10.5), and higher ionic strength up to 1.0 mol/L KCl promoted 20%-50% higher phosphate release.