Goormaghtigh E, Cabiaux V, Ruysschaert J M
Laboratoire de Chimie Physique des Macromolécules aux Interfaces, Université Libre de Bruxelles, Belgium.
Eur J Biochem. 1990 Oct 24;193(2):409-20. doi: 10.1111/j.1432-1033.1990.tb19354.x.
Attenuated total reflection Fourier-transform infrared spectroscopy of thin hydrated films of soluble and membrane protein included in a phospholipid bilayer is shown to provide useful information as to the secondary structure of the protein. The analysis of the amide I band of deuterated samples by Fourier self-deconvolution followed by a curve fitting was performed by a new procedure in which all the input parameters are generated by the computer rather than by the investigator. The results of this analysis provide a correct estimation of the alpha-helix and beta-sheet structure content with a standard deviation of 8.6% when X-ray structures are taken as a reference. We also show that the orientation of the different secondary structures resolved by the Fourier self-deconvolution/curve-fitting procedure and of the phospholipid acyl chains can be simultaneously evaluated for membrane proteins reconstituted in a lipid bilayer. Of special interest for reconstitution of membrane proteins, the lipid/protein ratio can be accurately and quickly determined from the infrared spectrum.
研究表明，对包含在磷脂双分子层中的可溶性蛋白和膜蛋白的薄水合膜进行衰减全反射傅里叶变换红外光谱分析，可为蛋白质的二级结构提供有用信息。通过傅里叶自卷积然后进行曲线拟合对氘代样品的酰胺I带进行分析，采用了一种新程序，其中所有输入参数均由计算机生成而非研究人员手动设定。以X射线结构为参考时，该分析结果能正确估计α-螺旋和β-折叠结构含量，标准差为8.6%。我们还表明，对于重构在脂质双分子层中的膜蛋白，通过傅里叶自卷积/曲线拟合程序解析的不同二级结构以及磷脂酰基链的取向可以同时进行评估。对于膜蛋白的重构特别有意义的是，可以从红外光谱中准确快速地确定脂质/蛋白质比率。
