Hallaway P E, Hallaway B E, Rosenberg A
Biochemistry. 1984 Jan 17;23(2):266-73. doi: 10.1021/bi00297a015.
We have studied the hydrogen-exchange kinetics of hemoglobin A0 as a function of ligand, CO, saturation. In the noncooperative system, azide binding to methemoglobin, the alterations in exchange kinetics are proportional to the average degree of ligation. However, in the case of CO binding to deoxyhemoglobin the changes in hydrogen-exchange pattern run ahead of the degree of ligation. The data can be best fitted assuming that all the liganded species, regardless of the number of ligands, show the same exchange properties. This two-state behavior must be the consequence of the fact that all the conformational changes leading to increased solvent accessibility take place when the first ligand is bound. Studies of the effect of pH changes and carbamoylation on the relationship between ligand binding and hydrogen exchange show that the observed differences of hydrogen exchange between deoxy and the liganded state are linked to the alkaline Bohr effect and to the state of the alpha-N-termini. As a consequence, at pH 9 despite a highly cooperative ligand binding isotherm the differences in hydrogen exchange between the deoxy and fully liganded species have vanished. We have to conclude that the hydrogen exchange is mirroring only the first part of the overall R to T transition. In all the experiments with pH changes and carbamoylation it is the liganded form that shows changes becoming more like the deoxy state. This is not consistent with a model where ligand binding removes a structural restriction in the less accessible deoxy state.
我们研究了血红蛋白A0的氢交换动力学与配体、一氧化碳（CO）饱和度的关系。在非协同体系中，叠氮化物与高铁血红蛋白结合时，交换动力学的改变与平均结合程度成正比。然而，在CO与脱氧血红蛋白结合的情况下，氢交换模式的变化比结合程度领先。假设所有结合配体的物种，无论配体数量如何，都表现出相同的交换特性，这些数据能得到最佳拟合。这种双态行为必定是由于当第一个配体结合时，所有导致溶剂可及性增加的构象变化都会发生这一事实。对pH变化和氨甲酰化对配体结合与氢交换关系的影响的研究表明，观察到的脱氧状态与结合配体状态之间氢交换的差异与碱性玻尔效应和α-N-末端的状态有关。因此，在pH 9时，尽管配体结合等温线具有高度协同性，但脱氧物种与完全结合配体的物种之间的氢交换差异已经消失。我们不得不得出结论，氢交换仅反映了从整体R态到T态转变的第一部分。在所有关于pH变化和氨甲酰化的实验中，都是结合配体的形式表现出变化，变得更像脱氧状态。这与一个模型不一致，在该模型中，配体结合消除了较难接近的脱氧状态下的结构限制。
