Evans J A, Gibbons I R
J Biol Chem. 1986 Oct 25;261(30):14044-8.
The presence of low concentrations of methanol or isopropyl alcohol (2-5%, v/v) in the assay medium stabilizes the latency of dynein 1 from sea urchin sperm flagella, with about a 50% decrease in ATPase level compared to that in the absence of solvent. Somewhat higher concentrations (10-20%, v/v) of these solvents in the assay give a 5-10-fold activation of ATPase activity. Dioxane, formamide, and dimethylformamide, on the other hand, always activate the ATPase activity, with a 5-10-fold increase observed at about 15% (v/v). The activation of latent ATPase activity by solvents is reversible for short exposures, especially in the presence of ATP and at low temperature, but the activation becomes irreversible upon more prolonged exposure. The rate constant for irreversible activation by 16% methanol at 21 degrees C is 0.08 min-1, compared to rates of 0.44 and 0.02 min-1 for activation by 0.05% Triton X-100 at 21 and 0 degree C, respectively. The slowness of this reversible activation induced by methanol and by Triton X-100 suggests that it is the result of large-scale conformational changes in the structure of the dynein. However, the activation by methanol occurs without the dissociation of the alpha and beta subunits of dynein that is observed with Triton X-100. The presence of 1 mM MgATP, or of 100 microM MgATP and 10 microM vanadate substantially protects latent dynein from activation by 0.05% Triton X-100.
测定介质中存在低浓度的甲醇或异丙醇（2 - 5%，v/v）可稳定海胆精子鞭毛中动力蛋白1的潜伏期，与不存在溶剂时相比，ATP酶水平降低约50%。在测定中，这些溶剂浓度稍高（10 - 20%，v/v）时，ATP酶活性会有5 - 10倍的激活。另一方面，二氧六环、甲酰胺和二甲基甲酰胺总是能激活ATP酶活性，在约15%（v/v）时可观察到活性增加5 - 10倍。溶剂对潜在ATP酶活性的激活在短时间暴露下是可逆的，特别是在有ATP存在且低温的情况下，但长时间暴露后激活会变为不可逆。21℃时16%甲醇不可逆激活的速率常数为0.08 min⁻¹，相比之下，0.05% Triton X - 100在21℃和0℃时激活的速率分别为0.44和0.02 min⁻¹。甲醇和Triton X - 100诱导的这种可逆激活较慢，表明这是动力蛋白结构发生大规模构象变化的结果。然而，甲醇激活时不会出现Triton X - 100激活时观察到的动力蛋白α和β亚基解离现象。1 mM MgATP或100 μM MgATP与10 μM钒酸盐的存在可显著保护潜在动力蛋白不被0.05% Triton X - 100激活。
