Okonjo K O, Vega-Catalan F J, Ubochi C I
Department of Chemistry, University of Ibadan, Nigeria.
J Mol Biol. 1989 Jul 20;208(2):347-54. doi: 10.1016/0022-2836(89)90394-x.
Temperature jumps on mixtures of hemoglobin and pH indicators give rise to relaxation signals in the microsecond range. The pH and concentration dependences of the reciprocal relaxation time, 1/tau, may be rationalized on the basis of a reaction scheme in which a slow isomerization process in the protein moiety is coupled to a rapid co-operative ionization of two protons. At 11 degrees C the rate constants of the isomerization are kr = 4.2(+/- 1.8) x 10(4) s-1 and kf = 1.3(+/- 0.1) x 10(4) s-1 in deoxyhemoglobin; in carbonmonoxyhemoglobin they are kr = 3.9(+/- 1.3) x 10(4) s-1 and kf = 5.3(+/- 1.8) x 10(3) s-1. The pKa values of the coupled ionizing groups are 5.3 in deoxy- and 6.0 in carbonmonoxyhemoglobin. Modification of the CysF9(93) beta sulfhydryl group with iodoacetamide abolishes the pH dependence of 1/tau, suggesting that this sulfhydryl is involved in the isomerization process. Consideration of the X-ray structure of oxyhemoglobin allows a structural interpretation of the results. It is concluded that the isomerization may be important for the physiological function of hemoglobin, but that it is not identical with the quaternary structure transition.
血红蛋白与pH指示剂混合物的温度跃升会产生微秒级的弛豫信号。基于一种反应机制，可对倒数弛豫时间1/τ的pH和浓度依赖性作出合理解释，该机制中蛋白质部分的缓慢异构化过程与两个质子的快速协同电离相耦合。在11摄氏度时，脱氧血红蛋白中异构化的速率常数为kr = 4.2(±1.8)×10⁴ s⁻¹和kf = 1.3(±0.1)×10⁴ s⁻¹；在碳氧血红蛋白中，它们分别为kr = 3.9(±1.3)×10⁴ s⁻¹和kf = 5.3(±1.8)×10³ s⁻¹。耦合电离基团的pKa值在脱氧血红蛋白中为5.3，在碳氧血红蛋白中为6.0。用碘乙酰胺修饰半胱氨酸F9(93)β巯基会消除1/τ对pH的依赖性，这表明该巯基参与了异构化过程。对氧合血红蛋白X射线结构的研究使得能够对结果进行结构解释。得出的结论是，异构化可能对血红蛋白的生理功能很重要，但它与四级结构转变并不相同。
