| The purpose of this thesis was to construct novel monolayer films of cytochrome c oxidase (CcO) with cytochrome c (cyt c) on surfaces such that the native properties of the proteins were retained and so the mechanism of catalytic action of the enzyme can be studied. The use of monolayer films and cyclic voltammetry (CV) to investigate redox proteins is a new methodology that can provide useful information about the coupling of electron transfer to proton transfer or other chemical events within enzymes and smaller proteins. However, there are considerable obstacles to overcome in the creation of viable protein films. Most importantly, adsorption to surfaces requires avoidance of the denaturing of vulnerable proteins such as cyt c, the natural C cO electron donating partner. Secondly, it is necessary to achieve a specific binding interaction with the surface to make an ordered film in which electron transfer to a large enzyme is sufficiently fast. In this respect, we used cyt c as a tether and natural bridge for communication with CcO in self-assembled films. This physiological arrangement led to the observation of substantial catalytic activity by the oxidoreductase with electrons donated from the electrode through cyt c, a previously unwitnessed event. In addition, a link with the past was established by performing optical spectroscopy and redox potentiometry on Langmuir-Blodgett monolayer films of CcO. These monolayers showed classic redox titrations of the two a-hemes in the enzyme and oxidation of cyt c from solution, proving retention of native characteristics in these films as well. Therefore, the stage was set for further electrometric and spectroscopic analysis of aspects of interest in the function of C cO. |
