Nanoparticulate glutathione peroxidase mimics based on selenocystine-pullulan conjugates.
ABSTRACT
We synthesized nanoparticulate glutathione peroxidase (GPx) mimics in which
selenocystine (SeCyst) was conjugated to a hydrophilic linear polysaccharide,
pullulan (Pul). The SeCyst ester-conjugated Pul derivatives (SeCyst-Pul)
in phosphate buffer (pH 7) were treated with a sonicator to spontaneously
form particulate materials. Dynamic light scattering measurements revealed
that the SeCyst-Pul conjugates could form particulate materials with diameters
between 100 and 300 nm. Distinctive endothermic peaks were observed for
the SeCyst-Pul aggregate solutions based on a differential scanning calorimetric
analysis. The tryptophan (Trp) fluorescence intensity of SeCyst benzyl
ester-tryptophanyl-Pul (SeCyst-Bz-Trp-Pul) mostly decreased in comparison
to those of the Trp-Pul (its precursor) and free Trp, which indicates that
the Trp residues come close to each other during the aggregation of the
conjugates. Formation of SeCyst-Pul aggregates could be induced by the
hydrophobic interactions between the SeCyst esters and the amino acid residues
on Pul. The GPx-like activity of SeCyst-Bz-Trp-Pul aggregates for the reduction
of H2O2 was enhanced nearly 20-fold higher than that of free SeCyst. The double-reciprocal
plots of the SeCyst-Bz-Trp-Pul aggregate-catalyzed reduction yielded parallel
lines by varying the substrate concentrations, indicating a "ping-pong"
mechanism that is similar to those of the natural GPxs. The enhanced GPx
activity of the SeCyst-Bz-Trp-Pul aggregate was also supported by higher
kinetic parameters, kcat/Km GSH and kcat/Km H2O2. Overall, the enhanced activity of the SeCyst-Bz-Trp-Pul aggregate would
be attributed to a hydrophobic environment that was formed at the vicinity
of the SeCyst.