(187af) Markedly Increased Activity of Lysozyme Adsorbed on SBA-15 Silica with Controlled Nanopore Diameter

Markedly Increased
Activity of Lysozyme Adsorbed on SBA-15 Silica with Controlled Nanopore
Diameter

With
their high surface area and well-controlled pore morphology, nanostructured mesoporous
materials are increasingly considered as supports for enzymes, for applications
in industrial catalysis, membranes and sensing. While research has mainly
focused on these applications, our fundamental understanding of the local
effects of a support on an enzyme remains unclear. This, in turn, is important
for the rational design of immobilized enzymes for targeted applications.
Answering the question of how the local curvature of nanostructured materials
influences enzyme activity is the objective of our research.

Here
we report how the local curvature of mesoporous materials affects the diffusion
and adsorption of lysozyme, and its catalytic activity, when it is immobilized
on mesoporous supports. Lysozyme is a test molecule in our fundamental study,
because it is almost spherical and many data have been reported in the
literature. As mesoporous supports, we synthesized a series of SBA-15s, which
are silicas with a well-defined, hexagonal array of parallel mesopores of the
same pore diameter. The pore diameter of SBA-15s was varied between 5 and 12 nm
by changing the synthesis conditions. Given the similar dimensions of the pores
and lysozyme (approximately 3.0 X 3.0 X 4.5 nm³), confinement
effects may occur. We measured the kinetics and the thermodynamics of lysozyme
adsorption on this series of SBA-15 experimentally, and compared the results to
model calculations, as a function of pore size and surface area.

The
surface of mesoporous materials might change the conformation of the attached
molecules, and their activity. When lysozyme was immobilize on to the internal
surface of SBA-15, whose local curvature changed with the pore diameter, we
demonstrated a dramatic increase in enzyme activity, for experiments with a
substrate (4-Methylumbelliferyl-N,N',N''-triacetyl-Beta-chitotrioside,
MW=787.75 g/mol) small enough to enter the pores. However, the activity when
using whole cells (Micrococcus lysodeikticus, D=0.8µm), which cannot
enter the nanopores, dropped to almost zero, proving that the enzyme is indeed
mostly adsorbed on the internal surface, and that it is this enzyme which is
responsible for the activity increase in the first experiments. Results with
external adsorption on nanoparticles and nanotubes by others [1, 2] had
indicated a constant or dropped activity, rather than an increase. This
suggests remarkable confinement effects in nanoporous media, induced by
geometrical and/or chemical effects, which further studies on other supports
and with different substrates will help to elucidate.

References:

[1]
Alexey A. Vertegel, Richard W. Siegel, Jonathan S. Dordick; Silica Nanoparticle
size influences the structure and enzymatic activity of adsorbed lysozyme; Langmuir
2004, 20, 6800-6807

[2]
Hao-Min Ding, Lei Shao, Run-Jing Liu, Qing-Gui Xiao, Jian-Feng Chen; Silica
nanotubes for lysozyme immobilization; J.  Coll. Inter. Sci. 2005,
290(1), 102-106