Gilbert J M, Strocchi P, Brown B A, Marotta C A
J Neurochem. 1981 Mar;36(3):839-46. doi: 10.1111/j.1471-4159.1981.tb01670.x.
Free and membrane-bound polysomes were prepared from rat forebrain and added to a cell-free system containing rabbit reticulocyte factors and L-[35S]methionine. The translation products were analyzed by two-dimensional gel electrophoresis followed by autoradiography. The free polysomes synthesized actin and at least four major tubulin subunits (alpha 1, alpha 2, beta 1, and beta 2) that are found in rat forebrain cytoplasm. The membrane-bound polysomes synthesized predominantly one protein (MB) in the tubulin region of the two-dimensional gel. MB has a molecular weight and isoelectric point similar to alpha-tubulin. Only trace amounts of alpha- and beta-tubulin and action were synthesized by the membrane-bound polysomes. MB co-purified with cytoplasmic tubulin after two cycles of aggregation and disaggregation. MB synthesized in vitro (from membrane-bound polysomes) and alpha- and beta-tubulin and actin subunits (synthesized from free polysomes) were digested with Staphylococcus aureus V8 protease, and the resulting peptides were separated by slab gel electrophoresis followed by autoradiography. The peptide pattern of MB was similar but not identical to the peptide patterns of alpha- and beta-tubulin; MB yielded peptides not found in tubulin. We conclude that membrane-bound polysomes from rat forebrain do not synthesize significant amounts of the predominant tubulin subunits synthesized by free polysomes. A major protein (MB) is synthesized by membrane-bound polysomes and is similar, but not identical, to alpha-tubulin synthesized by free polysomes on the basis of molecular weight, isoelectric point, and peptide analysis.
从大鼠前脑制备游离和膜结合多核糖体，并将其添加到含有兔网织红细胞因子和L-[35S]甲硫氨酸的无细胞系统中。通过二维凝胶电泳和放射自显影分析翻译产物。游离多核糖体合成肌动蛋白和至少四种主要的微管蛋白亚基（α1、α2、β1和β2），这些亚基存在于大鼠前脑细胞质中。膜结合多核糖体在二维凝胶的微管蛋白区域主要合成一种蛋白质（MB）。MB的分子量和等电点与α-微管蛋白相似。膜结合多核糖体仅合成微量的α-和β-微管蛋白以及肌动蛋白。经过两轮聚集和解聚后，MB与细胞质微管蛋白共纯化。用金黄色葡萄球菌V8蛋白酶消化体外合成的MB（来自膜结合多核糖体）以及α-和β-微管蛋白和肌动蛋白亚基（由游离多核糖体合成），所得肽段通过平板凝胶电泳和放射自显影分离。MB的肽谱与α-和β-微管蛋白的肽谱相似但不相同；MB产生了微管蛋白中未发现的肽段。我们得出结论，大鼠前脑的膜结合多核糖体不会合成游离多核糖体合成的大量主要微管蛋白亚基。膜结合多核糖体合成一种主要蛋白质（MB），基于分子量、等电点和肽分析，它与游离多核糖体合成的α-微管蛋白相似但不相同。
