Shapiro D B, Esquerra R M, Goldbeck R A, Ballas S K, Mohandas N, Kliger D S
Department of Chemistry and Biochemistry, University of California, Santa Cruz 95064, USA.
J Biol Chem. 1995 Nov 3;270(44):26078-85. doi: 10.1074/jbc.270.44.26078.
The re-equilibration rate of carbon monoxide binding to hemoglobin S polymers is determined by time-resolved measurements of linear dichroism spectra. Linear dichroism is used to detect religation to hemoglobin in the polymer in the presence of rebinding to free hemoglobin S tetramers. Measurement of the linear dichroism resulting from photolysis of the small percentage of ligand bound to the polymer is accomplished through the use of an ultrasensitive, ellipsometric linear dichroism technique developed for this purpose. The major finding is that the return of the polymer phase to its equilibrium ligation state is much slower than that of the solution phase hemoglobin tetramers. Assuming all hemes in the polymer are equally likely to participate in rebinding, the re-equilibration rate for carbon monoxide religation to hemoglobin S polymers is found to be 0.14 +/- 0.07 (s-1 mM-1), about 1000 times slower than the rebinding rate of carbon monoxide to T-state monomer hemoglobin. Several interpretations of this result are discussed. An understanding of the ligand binding kinetics to hemoglobin S polymers could have pathophysiological significance in its relevance to polymer formation and melting during red blood cell circulation.
通过对线性二色性光谱进行时间分辨测量来确定一氧化碳与血红蛋白S聚合物的重新平衡速率。在存在与游离血红蛋白S四聚体重新结合的情况下，线性二色性用于检测聚合物中血红蛋白的重新结合。通过使用为此目的开发的超灵敏椭圆偏振线性二色性技术，来测量与聚合物结合的小百分比配体光解所产生的线性二色性。主要发现是聚合物相恢复到其平衡结合状态的速度比溶液相血红蛋白四聚体慢得多。假设聚合物中的所有血红素参与重新结合的可能性相同，发现一氧化碳与血红蛋白S聚合物重新结合的重新平衡速率为0.14±0.07（s-1 mM-1），比一氧化碳与T态单体血红蛋白的重新结合速率慢约1000倍。讨论了该结果的几种解释。了解配体与血红蛋白S聚合物的结合动力学在其与红细胞循环过程中聚合物形成和熔化的相关性方面可能具有病理生理学意义。
