Ho C, Baldassare J J, Charache S
Proc Natl Acad Sci U S A. 1970 Jul;66(3):722-9. doi: 10.1073/pnas.66.3.722.
The spin label technique has been used to study human hemoglobins A, F, Zürich, and Chesapeake as a function of carbon monoxide saturation. The experimental results suggest that the changes in the electron paramagnetic resonance spectra of hemoglobin labeled with N-(1-oxyl-2,2,6,6-tetramethyl-4-piperidinyl)iodoacetamide depend on the state of ligation of more than one heme group. For those hemoglobins with full or large cooperative ligand binding (such as A, F, and Zürich), there is a lack of isosbestic points in the spectra as a function of CO saturation. However, for those hemoglobins with little or no cooperative ligand binding (such as Chesapeake and methemoglobins), there is a sharp set of isosbestic points. These findings confirm and extend the early work of McConnell and co-workers. The absence of a set of isosbestic points in those hemoglobins with full cooperative ligand binding is consistent with the sequential model of Koshland, Némethy, and Filmer for cooperative oxygen binding to hemoglobin. The present results, with hemoglobin variants having known amino acid substitutions, also focus on the importance of the interactions among the amino acid residues located at alpha(1)-beta(2) or alpha(2)-beta(1) subunit contacts for the functioning of hemoglobin as an oxygen carrier. In addition, the resonance spectra of the spin label are very sensitive to small structural variations around the heme groups in the beta- or gamma-chains where the labels are attached. The results of the spin label experiment are discussed in relation to recent findings on the mechanism of oxygenation of hemoglobin from the nuclear magnetic resonance studies of this laboratory and the x-ray crystallographic analysis of Perutz and co-workers.
自旋标记技术已被用于研究人类血红蛋白A、F、苏黎世型和切萨皮克型与一氧化碳饱和度的关系。实验结果表明，用N-(1-氧代-2,2,6,6-四甲基-4-哌啶基)碘乙酰胺标记的血红蛋白的电子顺磁共振光谱变化取决于不止一个血红素基团的连接状态。对于那些具有完全或高度协同配体结合的血红蛋白（如A、F和苏黎世型），光谱中缺乏作为一氧化碳饱和度函数的等吸收点。然而，对于那些具有很少或没有协同配体结合的血红蛋白（如切萨皮克型和高铁血红蛋白），有一组明显的等吸收点。这些发现证实并扩展了麦康奈尔及其同事的早期工作。那些具有完全协同配体结合的血红蛋白中缺乏一组等吸收点，这与科什兰德、内梅蒂和菲尔默提出的血红蛋白协同氧结合的顺序模型一致。目前使用已知氨基酸取代的血红蛋白变体的结果，也强调了位于α(1)-β(2)或α(2)-β(1)亚基接触处的氨基酸残基之间的相互作用对于血红蛋白作为氧载体发挥功能的重要性。此外，自旋标记的共振光谱对连接有标记的β链或γ链中血红素基团周围的小结构变化非常敏感。结合本实验室核磁共振研究和佩鲁茨及其同事的x射线晶体学分析中关于血红蛋白氧合机制的最新发现，讨论了自旋标记实验的结果。
