Flow microcalorimetric study of enzyme reactions: Application to arylesterase from human serum

The enzymatic hydrolysis of phenyl acetate, catalysed by arylesterase/paraoxonase (EC 3.1.8.1) was studied at 37 image C in Tris buffer, pH 8, by spectrophotometry and flow microcalorimetry, using an enzyme purified from human serum. After correction for buffer protonation and product ionization, the hydrolysis reaction was found to be slightly endothermic, with image kJ mol−1. Microcalorimetric data were analysed with the integrated Michaelis equation to give the kinetic parameters of the enzyme: Michaelis constant image mM, catalytic constant image s−1, bimolecular rate constant image M−1 s−1. These results were in agreement with the spectrophotometric method. This study confirms the usefulness of microcalorimetry in the field of enzyme kinetics.