Perrella M, Benazzi L, Shea M A, Ackers G K
Dipartimento di Scienze e Technologie Biomediche, Universita di Milano, Italy.
Biophys Chem. 1990 Jan;35(1):97-103. doi: 10.1016/0301-4622(90)80064-e.
Reaction of tetrameric hemoglobin with ligands at the four heme sites yields nine species that have structurally unique combinations of ligated and unligated subunits. Using hemoglobins where the ligated subunits contain cyanomethemoglobin, Smith and Ackers studied the dimer-tetramer assembly reactions in all nine of the partially ligated species (F. R. Smith and G. K. Ackers, Proc. Natl. Acad. Sci. U.S.A. 82 (1985) 5347). They found a third assembly free energy in addition to those of unligated hemoglobin and fully ligated cyanomethemoglobin. The observed distribution of the three assembly free energies among the ten species was found to be incompatible with the two-state mechanism of allosteric control (J. Monod, J. Wyman and J. P. Changeaux, J. Mol. Biol. 12 (1965) 81). The results indicated a mechanism of 'combinatorial switching' in which the binding free energies per site change with configuration of occupied sites and not just their number. In this study, we have confirmed the existence of three assembly free energies among the ten ligation species using a cryogenic method (M. Perrella and L. Rossi-Bernardi, Methods Enzymol. 76 (1981) 133). For one of the species we find a different free energy assignment from that reported by Smith and Ackers; for all other species we observe the same assignments as in earlier work. The revised distribution also requires a 'combinatorial' mechanism of allosteric switching among the three states.
四聚体血红蛋白在四个血红素位点与配体反应会产生九种物种，这些物种具有连接和未连接亚基的结构独特组合。史密斯和阿克斯使用连接亚基含有氰化高铁血红蛋白的血红蛋白，研究了所有九种部分连接物种中的二聚体 - 四聚体组装反应（F.R.史密斯和G.K.阿克斯，《美国国家科学院院刊》82 (1985) 5347）。他们发现除了未连接血红蛋白和完全连接氰化高铁血红蛋白的组装自由能之外，还有第三种组装自由能。发现这三种组装自由能在这十种物种中的观察分布与变构控制的两态机制不兼容（J.莫诺德、J.怀曼和J.P.尚热，《分子生物学杂志》12 (1965) 81）。结果表明存在一种“组合切换”机制，其中每个位点的结合自由能随占据位点的构型而变化，而不仅仅取决于其数量。在本研究中，我们使用低温方法（M.佩雷拉和L.罗西 - 贝尔纳迪，《酶学方法》76 (1981) 133）证实了十种连接物种中存在三种组装自由能。对于其中一种物种，我们发现其自由能分配与史密斯和阿克斯报道的不同；对于所有其他物种，我们观察到与早期工作相同的分配。修订后的分布也需要三种状态之间变构切换的“组合”机制。
