Modulation
of Environmental Dynamics at the Active Site and Activity of an Enzyme
under Nanoscopic Confinement: Subtilisin Carlsberg in Anionic AOT
Reverse Micelle
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Abstract
Hydration
dynamics plays a crucial role in determining the structure, function,
dynamics, and stability of an enzyme. These dynamics involve the trapped-water
motions within small distance along with the total protein dynamics.
However, the exact molecular basis for the induction of enzyme function
by water dynamics is still remain unclear. Here, we have studied both
enzymatic activity and environmental dynamics at the active site of
an enzyme, Subtilisin Carlsberg (SC), under confined environment of
the reverse micelle (RM) retaining the structural integrity of the
protein. Kinetic measurements show that enzymatic activity increases
with increasing the water content of the RM. The picosecond-resolved
fluorescence Stokes shift studies indicate faster hydration dynamics
at the active site of the enzyme with increasing the water content
in the RM (<i>w</i><sub>0</sub> values). Temperature-dependent
hydration dynamics studies demonstrate the increased flexibility of
the protein at higher temperature under confinement. From temperature-dependent
solvation dynamics study, we have also calculated the activation energy
that has to be overcome for full orientational freedom to the water
molecules from bound to free-state. The results presented here establish
a correlation between the enzymatic activity and dynamics of hydration
of the encapsulated protein SC in cell-like confined environment within
the structural integrity of the enzyme