A qualitative and quantitative analysis of the conformation of Langmuir-Blodgett (LB) dried films of cytochrome C on silicon wafers was performed by Fourier transform ir (FTIR) spectroscopy. A deconvolution procedure was applied to the amide I band analysis, in order to determine the percentage of the different secondary structures. Qualitative analysis was performed by examining difference spectra. Films obtained by spreading protein solutions at pH 7.4 and 1, dried at 25 and 100 degrees C, on silicon wafers were also examined in order to detect spectral components associated with denatured protein domains, and to compare them with cytochrome C LB films. FTIR spectroscopy showed that the following important changes characterise LB film spectra(a) the alpha-helix component is higher (its percentage is 57 and 54%) than the one estimated in dried film obtained by spreading the solutions at pH 7.4 on a silicon substrate (43%), (b) there is an increase in the intensity of bands attributed to protonated carboxy group bands, involved and not involved in the formation of hydrogen bonds, and a decrease in those attributed to deprotonated carboxy groups, (c) the intensity of several bands attributed to aromatic amino acids and aliphatic chains increases, and (d) bands due to O-H stretching vibrations of crystallization water are present. These conformational changes could be induced by protein-protein interaction caused by the close packing of molecules that occurs during LB film formation; it cannot be excluded that they may be accompanied by partial changes in the tertiary structure of the protein. A preferential orientation of protein molecules in LB films is also a possibility.

译文

通过傅立叶变换红外光谱 (FTIR) 对硅晶片上细胞色素C的Langmuir-Blodgett (LB) 干膜的构象进行了定性和定量分析。将反卷积程序应用于酰胺I带分析,以确定不同二级结构的百分比。通过检查差异光谱进行定性分析。还检查通过在25和100 ℃ 下将pH 7.4和1下的蛋白质溶液在硅晶片上扩散而获得的膜,以便检测与变性蛋白质结构域相关的光谱组分,并将它们与细胞色素C LB膜进行比较。FTIR光谱表明,下列重要变化表征了LB薄膜光谱 (a) α-螺旋组分比在硅衬底上通过在pH 7.4下扩散溶液而获得的干膜中估计的 α-螺旋组分更高 (其百分比为57和54%) (43%),(b) 归因于质子化羧基带的带强度增加,参与和不参与氢键的形成,归因于去质子化羧基的带强度降低,(c) 归因于芳香族氨基酸和脂肪族链的几个带强度增加,并且 (d) 存在由于结晶水的o-h拉伸振动引起的带。这些构象变化可能是由LB膜形成过程中发生的分子紧密堆积引起的蛋白质-蛋白质相互作用引起的; 不能排除它们可能伴随着蛋白质三级结构的部分变化。LB膜中蛋白质分子的优先取向也是可能的。

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