Ifflaender U, Markau K, Sund H
Eur J Biochem. 1975 Mar 17;52(2):211-20. doi: 10.1111/j.1432-1033.1975.tb03989.x.
The theoretical and experimental analysis of a reversible association-dissociation equilibrium between different proteins (mixed association) is described. The experiments were performed with glutamate dehydrogenases from beef and rat liver. These enzymes are different, especially with respect to their association behavior. The association constant of rat liver glutamate dehydrogenase has been determined by light-scattering measurements. Its value (1.3 x 10(-4) M(-1)) is much lower than that of the beef liver enzyme, but the difference in the free association energy is only 30%. Association between these two enzymes is observed, also employing light-scattering experiments. Theoretical curves for mixed associating systems have been calculated and by comparison with these curves the mixed association constant could be determined. Since the free association energy of the mixed association is very near to the arithmetic mean between the values for the pure enzymes, the association interactions appear to be additive. The model of an open association with a virial coefficient is also true for the rat enzyme and the mixed association. The ultracentrifuge data are also explained by the same model and yield a similar value for the mixed association constant. Differences in the enzyme kinetics are small, but a somewhat reduced lifetime of the ternary complexes with the coenzymes and with subs-rates or GTP can be concluded for the rat liver enzyme. The circular dichroism measurements indicate no significant difference in the dissociation constants of the nucleotides, but the different amplitudes of the ellipticity indicate small differences in the electrical environment of the active center.
本文描述了不同蛋白质之间可逆缔合 - 解离平衡（混合缔合）的理论与实验分析。实验使用了牛肉和大鼠肝脏中的谷氨酸脱氢酶。这些酶不同，尤其是在缔合行为方面。大鼠肝脏谷氨酸脱氢酶的缔合常数已通过光散射测量确定。其值（1.3×10⁻⁴ M⁻¹）远低于牛肉肝脏酶的值，但自由缔合能的差异仅为30%。通过光散射实验也观察到了这两种酶之间的缔合。计算了混合缔合系统的理论曲线，并通过与这些曲线比较确定了混合缔合常数。由于混合缔合的自由缔合能非常接近纯酶值的算术平均值，缔合相互作用似乎是可加的。具有维里系数的开放缔合模型对于大鼠酶和混合缔合也适用。超速离心数据也由同一模型解释，并得出了相似的混合缔合常数值。酶动力学的差异较小，但可以推断大鼠肝脏酶与辅酶以及与底物或GTP形成的三元复合物的寿命有所缩短。圆二色性测量表明核苷酸解离常数没有显著差异，但椭圆率的不同幅度表明活性中心的电环境存在微小差异。
