红外光谱酰胺Ⅲ带用于蛋白质二级结构的测定研究

Studies on Amide Ⅲ Infrared Bands for the Secondary Structure Determination of Proteins

用甲醇对 BSA和 Rase A等蛋白质进行变性处理 ,结合蛋白质酰胺 带的拟合结果对酰胺 带各二级结构的谱峰进行了初步指认 :1 330～ 1 2 90 cm- 1为 α-螺旋 ;1 2 95～ 1 2 6 5 cm- 1为 β-转角 ;1 2 70～ 1 2 4 5cm- 1 为无规卷曲 ;1 2 5 0～ 1 2 2 0 cm- 1 为β-折叠 .依据这些谱峰归属 ,对一些已知二级结构的蛋白质进行了测定 ,所得结果与 X射线衍射数据以及酰胺 带的定量结果基本一致 .

Fourier transform infrared spectroscopy is increasingly becoming an important method for quantitatively determining the secondary structure of proteins. Amide Ⅰ band(1 600-1 700 cm -1) and amide Ⅲ (1 220-1 330 cm -1) are two main bands for this purposes. Amide Ⅲ was neglected because of its relatively weak in signals, but there is not interference from water and water vapor vibration bands and it is more sensitive to the changes of protein secondary structure. In this paper, proteins BSA and RNase A were denatured by methanol. The component bands of secondary structure in amide Ⅲ were assigned by combining the quantitative results of amide Ⅰ band. α-Helix 1 330-1 290 cm -1; β-turn 1 295-1 265 cm -1; Random coil 1 270-1 245 cm -1 and β-sheet 1 250-1 220 cm -1. The overlapping area between the neighboring bands of different structures, such as 1 290-1 295 cm -1, 1 265-1 270 cm -1, 1 245-1 250 cm -1, can be determined according to quantitative results of amide Ⅰ band. By using above assignments, the quantitative analysis results of the proteins in which the secondary structures have been known were consistent with that of X-ray data and amide Ⅰ band.