Fig. 3.

Determination of COS kinetic parameters at pH 7.5 and 25 °C in 200 mM Tris–HCl and 500 mM NaCl. (A) Calorimetric initial rate measurements. The figure shows a typical thermogram obtained by ITC in which a COS solution (2.46 µM) was titrated with choline-O-sulfate (300 mM) (2×5 µL and then 18×10 µL). Each titration was done before a significant amount of choline-O-sulfate was hydrolyzed. (B) Michaelis–Menten plot for the choline-O-sulfate hydrolysis. Data was obtained by transforming the calorimetric data to initial rates using procedures reported in the literature (see Section 2). (C) and (D) Michaelis–Menten plot for the hydrolysis of pNPS and MUS respectively obtained by UV–vis spectrophotometry.