Runge Anne F, Mendes Sergio B, Saavedra S Scott
Department of Chemistry and College of Optical Sciences, University of Arizona, Tucson, Arizona 85721, USA.
J Phys Chem B. 2006 Apr 6;110(13):6732-9. doi: 10.1021/jp056049e.
The structure of solution adsorbed and microcontact printed (muCP) cytochrome c (cyt c) films on glass and indium tin oxide (ITO) was investigated using attenuated total reflectance (ATR) and total internal reflectance fluorescence (TIRF) spectroscopies to determine the orientation of the heme groups in the films. The second and fourth order parameters of the heme as well as information on the angle between the absorption and emission dipoles of the heme, gamma, were experimentally determined. The order parameters of the heme are related to the order parameters of the protein molecule using the known angle between the heme plane and the electrostatic dipole moment of the cyt c protein. The effect of the surface roughness of the substrates (glass and ITO) was also taken into account quantitatively using AFM data. Physically possible order parameters were obtained for the heme group in both solution adsorbed and muCP films, but not for the electrostatic dipole moment of the protein. In addition, the experimental values of {cos2 gamma} for immobilized zinc-substituted cyt c are greater than the values of {cos2 gamma} determined in viscous solutions, which could be an indication that the environment of the heme groups changes upon adsorption. The electron transfer behavior of solution adsorbed and muCP films on ITO, determined using electrochemical methods, is compared to their orientation distribution and surface coverage as determined by spectroscopic methods.
使用衰减全反射（ATR）和全内反射荧光（TIRF）光谱研究了吸附在玻璃和氧化铟锡（ITO）上的溶液以及微接触印刷（μCP）细胞色素c（cyt c）薄膜的结构，以确定薄膜中血红素基团的取向。通过实验确定了血红素的二阶和四阶参数以及血红素吸收和发射偶极子之间夹角γ的信息。利用血红素平面与cyt c蛋白静电偶极矩之间的已知夹角，将血红素的序参数与蛋白质分子的序参数相关联。还使用原子力显微镜（AFM）数据定量考虑了基底（玻璃和ITO）表面粗糙度的影响。在溶液吸附和μCP薄膜中均获得了血红素基团物理上可能的序参数，但未获得蛋白质静电偶极矩的序参数。此外，固定化锌取代细胞色素c的{cos2γ}实验值大于在粘性溶液中测定的{cos2γ}值，这可能表明血红素基团的环境在吸附时发生了变化。使用电化学方法测定了ITO上溶液吸附和μCP薄膜的电子转移行为，并将其与通过光谱方法确定的取向分布和表面覆盖率进行了比较。
