Direct electron transfer-type bioelectrocatalysis of FAD-dependent glucose dehydrogenase using porous gold electrodes and enzymatically implanted platinum nanoclusters.

The direct electron transfer (DET)-type bioelectrocatalysis of flavin adenine dinucleotide (FAD)-dependent glucose dehydrogenase (GDH) from Aspergillus terreus (AtGDH) was carried out using porous gold (Au) electrodes and enzymatically implanted platinum nanoclusters (PtNCs). The porous Au electrodes were prepared by anodization of planar Au electrodes in a phosphate buffer containing glucose as a reductant. Moreover, PtNCs were generated into AtGDH by an enzymatic reduction of hexachloroplatinate (IV) ion. The modification was confirmed by native polyacrylamide gel electrophoresis and sodium dodecyl sulfate polyacrylamide gel electrophoresis analyses. The AtGDH-adsorbed porous Au electrode showed a DET-type bioelectrocatalytic wave both in the presence and absence of PtNCs; however, the current density with PtNCs (~1 mA cm-2 at 0 V vs. Ag|AgCl|sat. KCl) was considerably higher than that without PtNCs. The kinetic and thermodynamic analysis of the steady-state catalytic wave indicated that inner PtNCs shortened the distance between the catalytic center of AtGDH (=FAD) and the conductive material, and improved the heterogeneous electron transfer kinetics between them.

Crystallographic analysis of FAD-dependent glucose dehydrogenase.

An FAD-dependent glucose dehydrogenase (GDH) from Aspergillus terreus was purified and crystallized at 293 K using the sitting-drop vapour-diffusion method. A data set was collected to a resolution of 1.6 Šfrom a single crystal at 100 K using a rotating-anode X-ray source. The crystal belonged to space group P21, with unit-cell parameters a = 56.56, b = 135.74, c = 74.13 Å, ß = 90.37°. The asymmetric unit contained two molecules of GDH. The Matthews coefficient was calculated to be 2.2 Å(3) Da(-1) and the solvent content was estimated to be 44%.