阿尔伯塔血红蛋白（α2β2(101) 谷氨酸→甘氨酸）的功能及亚基组装特性

Functional and subunit assembly properties of hemoglobin Alberta (alpha 2 beta 2(101) Glu----Gly).

作者信息

McDonald M J, Turci S M, Bleichman M, Stinson R A

出版信息

J Mol Biol. 1985 May 5;183(1):105-12. doi: 10.1016/0022-2836(85)90284-0.

Abstract

Hemoglobin Alberta has an amino acid substitution at position 101 (Glu----Gly), a residue involved in the alpha 1 beta 2 contact region of both the deoxy and oxy conformers of normal adult hemoglobin. Oxygen equilibrium measurements of stripped hemoglobin Alberta at 20 degrees C in the absence of phosphate revealed a high affinity (P50 = 0.75 mm Hg at pH 7), co-operative hemoglobin variant (n = 2.3 at pH 7) with a normal Bohr effect (- delta log P50/delta pH(7-8) = 0.65). The addition of inositol hexaphosphate resulted in a decrease in oxygen affinity (P50 = 8.2 mm Hg at pH 7), a slight increase in the value of n and an enhanced Bohr effect. Rapid mixing experiments reflected the equilibrium results. A rapid rate of carbon monoxide binding (l' = 7.0 X 10(5) M-1 S-1) and a slow rate of overall oxygen dissociation (k = 15 s-1) was seen at pH7 and 20 degrees C in the absence of phosphate. Under these experimental conditions the tetramer stability of liganded and unliganded hemoglobin Alberta was investigated by spectrophotometric kinetic techniques. The 4K4 value (the liganded tetramer-dimer equilibrium dissociation constant) for hemoglobin Alberta was found to be 0.83 X 10(-6) M compared to a 4K4 value for hemoglobin A of 2.3 X 10(-6) M, indicating that the Alberta tetramer was less dissociated into dimers than the tetramer of hemoglobin A. The values of 0K4 (the unliganded tetramer-dimer equilibrium dissociation constant) for hemoglobin Alberta and hemoglobin A were also measured and found to be 2.5 X 10(-8) M and 1.5 X 10(-10) M, respectively, demonstrating a greatly destabilized deoxyhemoglobin tetramer for hemoglobin Alberta compared to deoxyhemoglobin A. The functional and subunit dissociation properties of hemoglobin Alberta appear to be directly related to the dual role of the beta 101 residue in stabilizing the tetrameric form of the liganded structure, while concurrently destabilizing the unliganded tetramer molecule.

摘要

阿尔伯塔血红蛋白在第101位氨基酸发生了取代（谷氨酸→甘氨酸），该残基参与正常成人血红蛋白的脱氧和氧合构象的α1β2接触区域。在20℃且无磷酸盐存在的条件下，对脱辅基阿尔伯塔血红蛋白进行的氧平衡测量显示，它具有高亲和力（在pH 7时P50 = 0.75 mmHg），是一种具有协同性的血红蛋白变体（在pH 7时n = 2.3），且具有正常的玻尔效应（-Δlog P50/ΔpH(7 - 8) = 0.65）。添加肌醇六磷酸会导致氧亲和力降低（在pH 7时P50 = 8.2 mmHg），n值略有增加，玻尔效应增强。快速混合实验反映了平衡结果。在pH 7和20℃且无磷酸盐存在的条件下，观察到一氧化碳的快速结合速率（l' = 7.0×10⁵ M⁻¹ s⁻¹）和整体氧解离的缓慢速率（k = 15 s⁻¹）。在这些实验条件下，通过分光光度动力学技术研究了配体化和未配体化的阿尔伯塔血红蛋白的四聚体稳定性。发现阿尔伯塔血红蛋白的4K4值（配体化四聚体 - 二聚体平衡解离常数）为0.83×10⁻⁶ M，而血红蛋白A的4K⁴值为2.3×10⁻⁶ M，这表明阿尔伯塔四聚体比血红蛋白A的四聚体更不容易解离成二聚体。还测量了阿尔伯塔血红蛋白和血红蛋白A的0K4值（未配体化四聚体 - 二聚体平衡解离常数），分别为2.5×10⁻⁸ M和1.5×10⁻¹⁰ M，这表明与脱氧血红蛋白A相比，阿尔伯塔血红蛋白的脱氧血红蛋白四聚体稳定性大大降低。阿尔伯塔血红蛋白的功能和亚基解离特性似乎直接与β101残基在稳定配体化结构的四聚体形式同时使未配体化的四聚体分子不稳定方面的双重作用有关。