2 research outputs found
A Fluorescently Labeled Dendronized Polymer–Enzyme Conjugate Carrying Multiple Copies of Two Different Types of Active Enzymes
A hybrid structure of a synthetic dendronized polymer,
two different
types of enzymes (superoxide dismutase and horseradish peroxidase),
and a fluorescent dye (fluorescein) was synthesized. Thereby, a single
polymer chain carried multiple copies of the two enzymes and the fluorescein.
The entire attachment chemistry is based on UV/vis-quantifiable bis-aryl
hydrazone bond formation that allows direct quantification of bound
molecules: 60 superoxide dismutase, 120 horseradish peroxidase, and
20 fluorescein molecules on an average polymer chain of 2000 repeating
units. To obtain other enzyme ratios the experimental conditions were
altered accordingly. Moreover, it could be shown that both enzymes
remained fully active and catalyzed a two-step cascade reaction
Structure and Enzymatic Properties of Molecular Dendronized Polymer–Enzyme Conjugates and Their Entrapment inside Giant Vesicles
Macromolecular hybrid structures
were prepared in which two types
of enzymes, horseradish peroxidase (HRP) and bovine erythrocytes Cu,Zn-superoxide
dismutase (SOD), were linked to a fluorescently labeled, polycationic,
dendronized polymer (denpol). Two homologous denpols of first and
second generation were used and compared, and the activities of HRP
and SOD of the conjugates were measured in aqueous solution separately
and in combination. In the latter case the efficiency of the two enzymes
in catalyzing a two-step cascade reaction was evaluated. Both enzymes
in the two types of conjugates were highly active and comparable to
free enzymes, although the efficiency of the enzymes bound to the
second-generation denpol was significantly lower (up to a factor of
2) than the efficiency of HRP and SOD linked to the first-generation
denpol. Both conjugates were analyzed by atomic force microscopy (AFM),
confirming the expected increase in object size compared to free denpols
and demonstrating the presence of enzyme molecules localized along
the denpol chains. Finally, giant phospholipid vesicles with diameters
of up to about 20 μm containing in their aqueous interior pool
a first-generation denpol–HRP conjugate were prepared. The
HRP of the entrapped conjugate was shown to remain active toward externally
added, membrane-permeable substrates, an important prerequisite for
the development of vesicular multienzyme reaction systems