Transparent Gold as a Platform for Adsorbed Protein Spectroelectrochemistry: Investigation of Cytochrome c and Azurin
2016-02-21T16:02:58Z (GMT) by
The majority of protein spectroelectrochemical methods utilize a diffusing, chemical mediator to exchange electrons between the electrode and the protein. In such methods, electrochemical potential control is limited by mediator choice and its ability to interact with the protein of interest. We report an approach for unmediated, protein spectroelectrochemistry that overcomes this limitation by adsorbing protein directly to thiol self-assembled monolayer (SAM) modified, thin (10 nm), semitransparent gold. The viability of the method is demonstrated with two diverse and important redox proteins: cytochrome c and azurin. Fast, reversible electrochemical signals comparable to those previously reported for these proteins on ordinary disk gold electrodes were observed. Although the quantity of protein in a submonolayer adsorbed at an electrode is expected to be insufficient for detection of UV–vis absorption bands based on bulk extinction coefficients, excellent spectra were detected for each of the proteins in the adsorbed state. Furthermore, AFM imaging confirmed that only a single layer of protein was adsorbed to the electrode. We hypothesize that interaction of the relatively broad gold surface plasmon with the proteins’ electronic transitions results in surface signal enhancement of the molecular transitions of between 8 and 112 times, allowing detection of the proteins at much lower than expected concentrations. Since many other proteins are known to interact with gold SAMs and the technical requirements for implementation of these experiments are simple, this approach is expected to be very generally applicable to exploring mechanisms of redox proteins and enzymes as well as development of sensors and other redox protein based applications.