Development of a novel glucose enzyme fuel cell system employing protein engineered PQQ glucose dehydrogenase.

Glucose dehydrogenase harboring pyrroloquinoline quinone as the prosthetic group (PQQGDH) from Acinetobacter calcoaceticus is an ideal enzyme for the anode of biofuel cell, because of its oxygen insensitivity and high catalytic efficiency. However, the application of PQQGDH for the bioanode is inherently limited because of its instability. Using Ser415Cys mutant whose stability was greatly improved, we constructed the biofuel cell system employing the engineered PQQGDH as the bioanode enzyme and bilirubin oxidase (BOD) as the biocathode, and compared the stability of the biofuel cell with that employing wild-type PQQGDH. The maximum power density was 17.6 microW/cm2 at an external optimal load of 200 k omega. Using Ser415Cys mutant, the lifetime of the biofuel cell system was greatly extended to 152 h, more than six times as that of the biofuel cell employing the wild-type.

Glucose enzyme electrode using cytochrome b(562) as an electron mediator.

We demonstrate the construction of glucose sensors employing pyrroloquinoline quinone (PQQ) glucose dehydrogenase (PQQGDH) from Acinetobacter calcoaceticus and glucose oxidase (GOD) from Aspergillus nigar coupled with Escherichia coli soluble cytochrome b(562) (cyt b(562)) as electron acceptor. PQQGDH and GOD do not show direct electrochemical recycling of the prosthetic group at the electrode surface leading to a corresponding current signal. We constructed PQQGDH and GOD electrodes co-immobilized with 100-fold molar excess of cyt b(562) and investigated the electrochemical properties without synthetic electron mediators. PQQGDH/cyt b(562) and GOD/cyt b(562) electrodes both responded well to glucose whereas no current increase was observed from the electrode immobilizing enzyme alone. The detection limits for the PQQGDH/cyt b(562) and GOD/cyt b(562) electrodes were 0.1 and 0.8 mM, respectively, and their linearity extended to over 2 and 9 mM, respectively. These results demonstrate that a sensor system can be constructed without a synthetic electron mediator by using a natural electron acceptor. Furthermore, we have demonstrated the potential application of cyt b(562) in direct electron transfer type sensor systems with oxidoreductases whose quaternary structure do not contain any electron transfer subunit.