Yamamoto H, Saitoh Y, Yasugawa S, Miyamoto E
Department of Pharmacology, Kumamoto University Medical School, Japan.
J Neurochem. 1990 Aug;55(2):683-90. doi: 10.1111/j.1471-4159.1990.tb04187.x.
When the synaptosomal cytosol fraction from rat brain was chromatographed on a DEAE-cellulose column and assayed for protein phosphatases for tau factor and histone H1, two peaks of activities, termed peak 1 (major) and peak 2 (minor), were separated. Each peak was in a single form 2 (minor), were separated. Each peak was in a single form on Sephacryl S-300 column chromatography. Both peaks 1 and 2 dephosphorylated tau factor phosphorylated by Ca2+/calmodulin-dependent protein kinase II and the catalytic subunit of cyclic AMP-dependent protein kinase. The Km values were in the range of 0.42-0.84 microM for tau factor. There were no differences in kinetic properties of dephosphorylation between the substrates phosphorylated by the two kinases. The phosphatase activities did not depend on Ca2+, Mn2+ and Mg2+. Immunoprecipitation and immunoblotting analysis using polyclonal antibodies to the catalytic subunit of brain protein phosphatase 2A revealed that both protein phosphatases are the holoenzymic forms of protein phosphatase 2A. Aluminum chloride inhibited the activities of both peaks 1 and 2 with IC50 values of 40-60 microM. These results suggest that dephosphorylation of tau factor in presynaptic nerve terminals is controlled mainly by protein phosphatase 2A and that the neurotoxic effect of aluminum seems to be related mostly to inhibition of dephosphorylation of tau factor.
当将大鼠脑的突触体胞质溶胶部分在DEAE - 纤维素柱上进行层析，并针对tau因子和组蛋白H1的蛋白磷酸酶进行测定时，分离出了两个活性峰，分别称为峰1（主要）和峰2（次要）。在Sephacryl S - 300柱层析上，每个峰均呈现单一形式。峰1和峰2都能使由Ca2+/钙调蛋白依赖性蛋白激酶II和环磷酸腺苷依赖性蛋白激酶催化亚基磷酸化的tau因子去磷酸化。tau因子的米氏常数在0.42 - 0.84微摩尔范围内。两种激酶磷酸化的底物在去磷酸化的动力学性质上没有差异。磷酸酶活性不依赖于Ca2+、Mn2+和Mg2+。使用针对脑蛋白磷酸酶2A催化亚基的多克隆抗体进行免疫沉淀和免疫印迹分析表明，这两种蛋白磷酸酶均为蛋白磷酸酶2A的全酶形式。氯化铝抑制峰1和峰2的活性，半数抑制浓度（IC50）值为40 - 60微摩尔。这些结果表明，突触前神经末梢中tau因子的去磷酸化主要由蛋白磷酸酶2A控制，并且铝的神经毒性作用似乎主要与抑制tau因子的去磷酸化有关。
