Kinetic and thermodynamic properties of pseudomonas fluorescence lipase upon addition of proline.

The effect of proline on refolding and unfolding kinetics as well as activity of lipase from Pseudomonas fluorescens was determined using stopped-flow fluorescence and UV/Vis absorbance spectroscopy. Enzyme assay at different concentrations of proline revealed that activity of enzyme reaches maximum at 0.6 M concentration of proline. Kinetic measurements showed that refolding rate is considerably accelerated in the presence of 0.6M proline. Unfolding kinetic traces were fitted to double exponential function, and it was revealed that lipase molecules were unfolded via two different pathways. Fast unfolding rate constant decreased, while slow one did not change significantly upon addition of proline.