To investigate the structure of native Cyt c in solution we performed CD analysis at two different concentrations, 20 µM and 1 µM, to determine whether the intensity of maxima and minima peaks changed as the signal-to-noise ratio changed (lower Cyt c concentration should have a larger signal-to-noise ratio) (Figure ). The higher concentration of Cyt c was used to obtain an accurate CD spectrum, while the lower concentration was employed to represent the amount of Cyt c that is usually conjugated to AuNPs. In the far-UV CD spectra (in the range of 180-260 nm), the double minima in molecular ellipticity located at 208 and 222 nm and the positive band at ~190 nm can be identified for Cyt c Ox and Red in both 20 µM (Figure ) and 1 µM (Figure ). The double minima and positive band indicate the alpha-helical content, which can be associated with the catalytic ability of the protein . The double minima are more defined for Cyt c Red than Cyt c Ox in both Figure and Figure , insinuating a greater alpha helical content. The near-UV CD spectra in the range of 260-360 nm showed a double minimum at 282 nm 288 nm for both Cyt c Ox and Red at 20 µM (Figure ), which can be associated with Tryptophan residues indicative of the tertiary structure . Only the minimum at 288 nm is visible in Cyt c Ox and Red at 1 µM, with the 282 nm band appearing to blue shift to ~275 nm (Figure ), as the signalto-noise ratio must be large to detect this minimum at 282 nm. The CD spectra were also obtained in the Soret region (360-450 nm), enabling further understanding of the tertiary structure of Cyt c, as the Soret region can be attributed to the heme ring of Cyt c (Figure and). The 20 µM Cyt c Ox and Red showed maxima at 405 and 408 nm, respectively, with minima both at 416 nm. 20