Using Scanning Electrochemical Microscopy (SECM) to Measure the Electron-Transfer Kinetics of Cytochrome c Immobilized on a COOH-Terminated Alkanethiol Monolayer on a Gold Electrode

• Published in: Langmuir Vol. 22; no. 9; pp. 4298 - 4304
• Main Author: Holt, Katherine B
• Format: Journal Article
• Language: English
• Published: Washington, DC American Chemical Society 25.04.2006
• Subjects: Alkanes - chemistry; Animals; Chemistry; Cytochromes c - metabolism; Electrochemistry - methods; Electron Transport; Enzymes, Immobilized - metabolism; Exact sciences and technology; General and physical chemistry; Gold; In Vitro Techniques; Kinetics; Microelectrodes; Microscopy - methods; Models, Biological; Oxidation-Reduction; Sulfhydryl Compounds - chemistry; Binding; Self assembly; Monolayer; Gold; Immobilization; Mobility; Electron transfer; Alkanethiol; Transition metal; Potential; Orientation; Protein; Cytochrome c; Electrodes; Substrate; Ionic strength; Electrochemistry; Kinetics
• ISSN: 0743-7463; 1520-5827
• DOI: 10.1021/la0529916

Cytochrome c was electrostatically immobilized onto a COOH-terminated alkanethiol self-assembled monolayer (SAM) on a gold electrode at ionic strengths of less than 40 mM. Scanning electrochemical microscopy (SECM) was used to simultaneously measure the electron transfer (ET) kinetics of the bimolecular ET between a solution-based redox mediator and the immobilized protein and the tunneling ET between the protein and the underlying gold electrode. Approach curves were recorded with ferrocyanide as a mediator at different coverages of cytochrome c and at different substrate potentials, allowing the measurement of k BI = 2 × 108 mol-1 cm3 s-1 for the bimolecular ET and k° = 15 s-1 for the tunneling ET. The kinetics of ET was also found to depend on the immobilization conditions of cytochrome c:  covalent attachment gave slightly slower tunneling ET values, and a mixed CH3/COOH-terminated ML gave faster tunneling ET rates. This is consistent with previous studies and is believed to be related to the degree of mobility of cyt c in its binding configuration and its orientation with respect to the underlying electrode surface.