Electrical Contacting of an Assembly of Pseudoazurin and Nitrite Reductase Using DNA-Directed Immobilization

摘要

Abstract: A method for the electrical contacting of redox enzymes that obtain oxidizing or reducing equivalentsnfrom small electron-transfer proteins is demonstrated. The electrochemical contacting of redox enzymes throughntheir immobilization onto electrode supports offers great potential for technological applications and fornfundamental studies, but finding appropriate methods to immobilize the enzymes in an orientation allowingnrapid electron transfer with the electrode has proven difficult. The copper enzyme nitrite reductase (NiR) and itsnnatural electron-exchange partner pseudoazurin (Paz) are conjugated to a specific DNA tag and immobilizednto a gold electrode into a stoichiometrically defined assembly. The DNA tethered to the electrode surface actsnas flexible place-holder for the protein components, allowing both proteins to move within the construct. It isnshown that Paz efficiently shuttles electrons between the electrode and the NiR enzyme, allowing thenelectrochemically driven NiR catalysis to be monitored. The activity of the NiR enzyme remains unperturbed bynthe immobilization. The rate-limiting step of the system is tentatively ascribed to the dissociation of the Paz/NiRncomplex. The electrochemical response of the system reports not only on the NiR catalysis and on interfacialnelectron transfer but also on the interaction between NiR and Paz.