反相液相色谱中温度对蛋白分子构象变化的新表征

A New Characterization Parameter for Molecular Conformation Change of Proteins in Reversed-phase Liquid Chromatography

液相色谱中溶质计量置换保留模型的Ｚ值可用来对反相高效液相色谱中热变蛋白的分子构象进行表征。发现在异丙醇－甲酸４４％（Ｖ／Ｖ）－水体系为流动相时完全变性蛋白的Ｚ值随温度升高而降低，且Ｚ值与绝对温度倒数（１／Ｔ）成正比，因此蛋白分子构象变化可用Ｚ对１／Ｔ作图所得两条直线的斜率差值来表征，差值愈大，蛋白分子构象变化愈甚。该两直线的交点即为蛋白分子构象的突变温度。与通常采用的１ｇｋ＇对１／Ｔ的作图相比，用Ｚ对１／Ｔ作图对蛋白分子构象进行表征不仅对各种热变蛋白都有效，而且结果准确。

The Z value of the stoichiometric displacement model for retention can be used to be a new chracterization parameter for the molecular conformation of biopolymers caused by temperature in reversed-phase liquid chromatography(RPLC).It was found that when 2-propanol 44%(V/V)-formic acid-water was selected as a mobile phase,the Z values of four kinds of completely denaturated proteins decreased with the increase of temperature and the Z value changes would be directly proportional to the reciprocal of absolute termperature (1/T).The chracterization of the change in the molecular conformation of biopolymers in the instance can be obtained by means of the difference between the two slopes of each proteins from the plots of Z vs. 1/T,and the larger the difference,the greater the change in the molecular conformation of the protein is. The abrupt change temperatures for each proteins can be calculated from the intersect of the two straight lines. Compared to the characterization of molecular conformation of proteins with the plot of 1gk'(k'is the capacity factor)versus 1/T,the plot Z versus 1/T is not only valid for each type of proteins, but also is more accurate.