SINGLE ENZYME NANOPARTICLES

Enzymes are extremely useful in many industrial and pharmaceutical areas due to their ability
to catalyze reactions with high selectivity. Enzymes are being used for various applications in
energy and environment including carbon capture and sequestration (CCS). There are a wide
range of applications

 Biosensors
 Bioremediation
 Enzymatic synthesis in pharmaceutical and food industries
 Detergent industry

In order to extend their lifetime, significant efforts have been made to increase their stability
using protein as well as by chemical modification. Many researchers have explored the
immobilization of enzymes onto carriers, or entrapment within a matrix, framework or
nanoparticle with the hope of constricting the movement of the enzyme and shielding it from
aggressive environments, thus delaying the denaturation. Single enzyme nanoparticles
(SENs) are a state of art technology that allows immobilisation of individual enzyme
molecules in a nanometre scale cross-linked polymer matrix that is covalently bonded to the
enzyme. In most cases, multiple enzymes will be contained within a single nanoparticle or
matrix but in recent years researchers have begun to wrap up individual enzymes within
single enzyme nanoparticles (SENs). In these nanoparticles the enzyme is stabilized by a thin
shell, typically a polymer, prepared either by in situ polymerization from the enzyme surface
or by assembling a preformed polymer around it. Porous materials can afford high enzyme
loading but suffer a much greater diffusional limitation of substrate. For example, the value
of effectiveness factor η for α-chymotrypsin (CT) was reported to be ≈ 0.3 when it is
entrapped in polyacrylamide hydrogel below 0.1 when incorporated into hydrophobic plastics
and less than 103 when cross-linked. TEM Images of Single enzyme nanoparticles containing
α chymotrypsin (CT)showed that

 Hollow center matches the size and shape of CT.

 The dark image surrounding CT contains silicon (Energy Dispersive X-ray Analysis)

It was also found that Half-life at 30ºC was up to 143 days and No activity decrease at 4ºC
for five months.
Reduction in the size of enzyme-carrier materials can generally improve the efficiency of
immobilized enzymes. In the case of surface attachment, smaller particles can provide a
larger surface area for the attachment of enzymes leading to higher enzyme loading per unit
mass of particles.

Enzymes are being used for various applications in energy and environment including carbon
capture and sequestration (CCS).The amount of CO2 in the atmosphere is increasing.
Therefore, there is an urgent need to devise some strategies for sequestration of CO2. With
the objective of further improving the stability of the enzyme we report here a unique
protocol for the synthesis of SEN-CA (single enzyme nanoparticle (SEN) by modifying the
surface of carbonic anhydrase (CA) with a thin layer of organic/inorganic hybrid polymer)by
caging each enzyme molecule with a hybrid organic/inorganic biopolymer silica network by
using biopolymer such as chitosan. Individual enzyme molecules are stabilized within a
biopolymeric silica network of nanometer- scale thickness. A key result is that stabilization of
the activity was achieved with minimal substrate mass-transfer limitation compared to CA
entrapped in larger scale materials.

The use of magnetic nanoparticles for the enzyme immobilization has been reported and it
showed a good enzyme stabilization with covalently attached lipase on the magnetic Fe2O3
nanoparticles. SENs are an active and stable form of enzyme system. Since SENs are still
nanometer scale (less than 10 nm in size for the case of SEN-CT), they can be further
immobilized into mesoporous materials Since mesoporous materials provide a high surface
area, the immobilization of SENs in mesoporous silica would result in good volumetric
enzyme activity. We anticipate that the combination of SENs (active and stable form of
enzyme) and mesoporous materials (immobilization substrate with a large surface area and
tunable pore size) will result in an ideal enzyme system for various applications in
bioremediation, biosensors, and other biocatalytic processes.

SUBMITTED BY

ALONA SARA SAJAN

S2 MTECH EE

ROLL NO :01