Overexpression, purification, functional activity and pH dependent structural changes of EhPSAT. (A) SDS-PAGE analysis of E coli lysate over expressing EhPSAT and the purified protein. Lanes 1-4 represent molecular weight markers, uninduced culture, induced culture and purified protein, respectively. (B) pH-dependent enzymatic activity profile of EhPSAT. The data has been represented as percentage relative activity with highest activity observed at pH 8.5 taken as 100%, each point representing mean ± SD of three independent measurements. (C) pH induced changes in the secondary structure of EhPSAT. The effect of pH on the CD signal at 222 nm. The inset shows far-UV CD spectra at pH 6 (solid line), 7 (dashed line), 8 (dotted line) and 9 (dash-dotted line), respectively. (D) and (E) pH-induced changes in PyP and Trp fluorescence polarization, respectively. In both the panels the filled and the open symbols represent protein samples in absence and presence of 200 mM NaCl, respectively. (F) pH induced changes in fluorescence emission spectra of EhPSAT excited at 295 nm. The curves 1-4 represent protein samples incubated at pH 6, 7, 8 and 9, respectively.