Protein adsorption on nanoporous TiO2 films: a novel approach to studying photoinduced protein/electrode transfer reactions.
We have investigated the use of nanoporous TiO2 films as substrates for protein immobilisation. Such films are of interest due to their high surface area, optical transparency, electrochemical activity and ease of fabrication. These films moreover allow detailed spectroscopic study of protein/electrode electron transfer processes. We find that protein immobilisation on such films may be readily achieved from aqueous solutions at 4 degrees C with a high binding stability and no detectable protein denaturation. The nanoporous structure of the film greatly enhances the active surface area available for protein binding (by a factor of up to 850 for an 8 microns thick film). We demonstrate that the redox state of proteins such as immobilised cytochrome-c (Cyt-c) and haemoglobin (Hb) may be modulated by the application of an electrical bias potential to the TiO2 film, without the addition of electron transfer mediators. The binding of Cyt-c on the TiO2 films is investigated as a function of film thickness, protein concentration, protein surface charge and ionic strength. We demonstrate the potential use of immobilised Hb on such TiO2 films for the detection of dissolved CO in aqueous solutions. We further show that protein/electrode electron transfer may be initiated by UV bandgap excitation of the TiO2 electrode. Both photooxidation and photoreduction of the immobilised proteins can be achieved. By employing pulsed UV laser excitation, the interfacial electron transfer kinetics can be monitored by transient optical spectroscopy, providing a novel probe of protein/electrode electron transfer kinetics. We conclude that nanoporous TiO2 films may be useful both for basic studies of protein/electrode interactions and for the development of novel bioanalytical devices such as biosensors.
Topoglidis
E
E
Departments of Biochemistry and Chemistry, Imperial College of Science, Technology and Medicine, South Kensington, London, UK SW7 2AY.
Lutz
T
T
Willis
R L
RL
Barnett
C J
CJ
Cass
A E
AE
Durrant
J R
JR
D000327:Adsorption; D004566:Electrodes; D010084:Oxidation-Reduction; D010777:Photochemistry; D011506:Proteins / Q000737:chemistry*; D013499:Surface Properties; D014025:Titanium / Q000737:chemistry*
D011506:Proteins; C009495:titanium dioxide; D014025:Titanium