Cytochrome c oxidase (CcO) from Paracoccus denitfificans was immobilized in a strict orientation via a his-tag attached to subunit I on a gold film and reconstituted in situ into a protein-tethered bilayer lipid membrane. In this orientation, the cytochrome c (cyt c) binding site is directed away from the electrode pointing to the outer side of the protein-tethered bilayer lipid membrane architecture. The CcO can thus be activated by cyt c under aerobic conditions. Catalytic activity was monitored by impedance spectroscopy, as well as cyclic voltammetry. Cathodic and anodic currents of the CcO with cyt c added to the bulk solution were shown to increase under aerobic compared to anaerobic conditions. Catalytic activity was considered in terms of repeated electrochemical oxidation/reduction of the CcO/cyt c complex in the presence of oxygen. The communication of cyt c bound to the CcO with the electrode is discussed in terms of a hopping mechanism through the redox sites of the enzyme. Simulations supporting this hypothesis are included.
In situ monitoring of the catalytic activity of cytochrome c oxidase in a biomimetic architecture / Marcel G., Friedrich; Markus A., Plum; Santonicola, Mariagabriella; Vinzenz U., Kirste; Wolfgang, Knoll; Bernd, Ludwig; R. L. C., Naumann. - In: BIOPHYSICAL JOURNAL. - ISSN 0006-3495. - STAMPA. - 95:3(2008), pp. 1500-1510. [10.1529/biophysj.107.122747]
In situ monitoring of the catalytic activity of cytochrome c oxidase in a biomimetic architecture
SANTONICOLA, MARIAGABRIELLA;
2008
Abstract
Cytochrome c oxidase (CcO) from Paracoccus denitfificans was immobilized in a strict orientation via a his-tag attached to subunit I on a gold film and reconstituted in situ into a protein-tethered bilayer lipid membrane. In this orientation, the cytochrome c (cyt c) binding site is directed away from the electrode pointing to the outer side of the protein-tethered bilayer lipid membrane architecture. The CcO can thus be activated by cyt c under aerobic conditions. Catalytic activity was monitored by impedance spectroscopy, as well as cyclic voltammetry. Cathodic and anodic currents of the CcO with cyt c added to the bulk solution were shown to increase under aerobic compared to anaerobic conditions. Catalytic activity was considered in terms of repeated electrochemical oxidation/reduction of the CcO/cyt c complex in the presence of oxygen. The communication of cyt c bound to the CcO with the electrode is discussed in terms of a hopping mechanism through the redox sites of the enzyme. Simulations supporting this hypothesis are included.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.