Article ID: 23-68127
Direct electron transfer (DET)-enabled oxidoreductase has been utilized to develop mediator-free bioelectronic devices. Fungal pyrroloquinoline quinone-dependent dehydrogenase from Coprinopsis cinerea is an attractive quinohemoprotein in which the catalytic PQQ domain and cytochrome domain perform DET. Here, we examined the difference in the distance from the active center to the protein surface between PQQ and heme. Then, we studied the direct bioelectrocatalysis of the full-length enzyme by regulating the distance between the enzyme and electrode with various alkyl chains of self-assembled monolayers (SAMs). The catalytic currents by the full-length enzyme were obtained on SAM of chain alkyl lengths of C6 or more, while no catalytic currents have been obtained by the isolated PQQ domain. After approximately 15 Å from PQQ in the active site to the electrode surface, the onset potential of the current shifted from the redox potential of PQQred/PQQsemi to Hemered/Hemeox. The results indicated that the chain length-dependent electron transfer pathway from the PQQ domain directly to the electrode changed via the cytochrome domain. The amount of electroactive cytochrome domain that had immobilized decreased with increasing pH (pH 6.0 to pH 9.5). In direct bioelectrocatalysis through interdomain electron transfer of the cytochrome domain, kcat was found to be pH dependent with an optimal pH of 8.5; therefore, the rate-limiting step that governs pH dependence is likely the IET process.
