Greene L E, Eisenberg E
J Biol Chem. 1980 Jan 25;255(2):549-54.
The binding of heavy meromyosin (HMM), a soluble two-headed fragment of myosin, to F-actin was examined at mu = 0.22 M, 22 degrees C. The actin-HMM association constant was determined by having HMM and subfragment 1 (S-1) compete for sites on F-actin. In these experiments, varying concentrations of S-1 were added to a fixed concentration of HMM and F-actin. F-actin and bound fragments (HMM and S-1) then were sedimented and the concentration of unbound fragments was determined. The data were analyzed using a set of theoretical equations proposed by Hill ((1978) Nature 274, 825-826) that provide a simple way of analyzing the relative binding of one- and two-headed ligands. Using these equations, the actin-HMM association constant was determined to be 3 x 10(9) M-1, while under the same conditions the actin-S-1 association constant is 5 x 10(6) M-1 (determined in preceding paper (Greene, L. E., and Eisenberg, E. (1980) J. Biol, Chem. 255, 543-548)). Therefore, under these conditions, HMM binds 600-fold stronger to actin than does S-1, indicating that both of the HMM heads can bind strongly to actin.
在μ = 0.22 M、22℃的条件下，研究了肌球蛋白的可溶性双头片段重酶解肌球蛋白（HMM）与F-肌动蛋白的结合。通过让HMM和亚片段1（S-1）竞争F-肌动蛋白上的位点来测定肌动蛋白-HMM的结合常数。在这些实验中，将不同浓度的S-1添加到固定浓度的HMM和F-肌动蛋白中。然后使F-肌动蛋白和结合片段（HMM和S-1）沉淀，并测定未结合片段的浓度。使用Hill提出的一组理论方程（（1978年）《自然》274, 825 - 826）对数据进行分析，这些方程提供了一种分析单头和双头配体相对结合的简单方法。使用这些方程，测定肌动蛋白-HMM的结合常数为3×10⁹ M⁻¹，而在相同条件下肌动蛋白-S-1的结合常数为5×10⁶ M⁻¹（在前一篇论文中测定（Greene, L. E., and Eisenberg, E. (1980) J. Biol, Chem. 255, 543 - 548））。因此，在这些条件下，HMM与肌动蛋白的结合力比S-1强600倍，这表明HMM的两个头部都能与肌动蛋白紧密结合。
