DeArmond S J, Fajardo M, Naughton S A, Eng L F
Brain Res. 1983 Mar 7;262(2):275-82. doi: 10.1016/0006-8993(83)91018-1.
In situ and in vitro degradation of glial fibrillary acidic (GFA) protein in mouse spinal cord was examined with electroblots stained for GFA protein by the peroxidase anti-peroxidase method. Non-degraded, intact GFA protein had a molecular weight of 48 Kdaltons and isoelectric points ranging from pH 5.8 to 6.4. The molecular weights of immunoreactive degradation products ranged from 47 to 28 Kdaltons. All of the degradation products had acid shifted isoelectric points (pH 5.8-5.2). Degradation was prevented by chelating calcium with EGTA. In contrast to in situ degradation, degradation in vitro with 3 mM CaCl2 occurred at a faster rate. The effect of pH and temperature on the degradation process were determined by incubating homogenized spinal cords in 3 mM CaCl2 solutions varying in pH from 4 to 10 and at 4, 37, and 60 degrees C. The greatest number of immunoreactive bands with the lowest molecular weights occurred at pH 8 and 37 degrees C. The results suggest that turnover of glial filaments is in part controlled by a calcium dependent proteinase active near neutral pH similar to that postulated for neurofilament turnover.
采用过氧化物酶抗过氧化物酶法对胶质纤维酸性蛋白（GFA）进行电印迹染色，检测小鼠脊髓中GFA蛋白的原位和体外降解情况。未降解的完整GFA蛋白分子量为48千道尔顿，等电点范围为pH 5.8至6.4。免疫反应性降解产物的分子量范围为47至28千道尔顿。所有降解产物的等电点均向酸性偏移（pH 5.8 - 5.2）。用乙二醇双四乙酸（EGTA）螯合钙可防止降解。与原位降解相反，在3 mM氯化钙存在下的体外降解速率更快。通过在pH值为4至10、温度为4℃、37℃和60℃的3 mM氯化钙溶液中孵育匀浆脊髓，确定了pH值和温度对降解过程的影响。在pH 8和37℃时出现了分子量最低且免疫反应条带数量最多的情况。结果表明，胶质细丝的周转部分受一种钙依赖性蛋白酶控制，该酶在接近中性pH时具有活性，类似于神经细丝周转所假定的情况。
