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Investigations on the Q and CT Bands of Cytochrome c Submonolayer Adsorbed on an Alumina Surface Using Broadband Spectroscopy with Single-Mode Integrated Optical Waveguides

Department of Physics and Astronomy, University of Louisville, Louisville, Kentucky 40292, Department of Chemistry, Butler University, 4600 Sunset Avenue, Indianapolis, Indiana 46208.
The Journal of Physical Chemistry C (Impact Factor: 4.84). 05/2009; 113(19):8306-8312. DOI: 10.1021/jp810845e
Source: PubMed

ABSTRACT In this work, we report experimental results on the molar absorptivity of cytochrome c adsorbed at different submonolayer levels onto an aluminum oxide waveguide surface; our data show a clear dependence of the protein optical properties on its surface density. The measurements were performed using the broadband, single-mode, integrated optical waveguide spectroscopic technique, which is an extremely sensitive tool able to reach submonolayer levels of detection required for this type of studies. This investigation focuses on the molar absorptivity at the Q-band (centered at 525 nm) and, for the first time to our knowledge, the weak charge transfer (CT) band (centered at 695 nm) of surface-adsorbed cyt c. Polarized light in the spectral region from 450 to 775 nm was all-coupled into an alumina thin film, which functioned as a single-mode planar optical waveguide. The alumina thin-film waveguide used for this work had a thickness of 180 nm and was deposited on a glass substrate by the atomic layer deposition process. The protein submonolayer was formed on the alumina waveguide surface through electrostatic adsorption from an aqueous buffer solution at neutral pH. The optical properties of the surface-adsorbed cyt c were investigated for bulk protein concentrations ranging from 5 nM to 8200 nM in the aqueous buffer solution. For a protein surface density of 2.3 pmol/cm(2), the molar absorptivity measured at the charge transfer band was 335 M(-1) cm(-1), and for a surface density of 15 pmol/cm(2) was 720 M(-1) cm(-1), which is much closer to the value of cyt c dissolved in an aqueous neutral buffer (830 M(-1) cm(-1)). The modification of the protein molar absorptivity and its dependence on the surface density can most likely be attributed to conformational changes of the surface-adsorbed species.

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