Takahashi Y, Ogata K
Biochim Biophys Acta. 1982 Apr 26;697(1):101-12. doi: 10.1016/0167-4781(82)90050-1.
The inhibitory effects of ethionine treatment of female rats for 4 h on the protein-synthesizing machineries of 80 S ribosomes and 40 S ribosomal subunits of the liver were investigated. The following results were obtained. (1) The translation of globin mRNA by 80 S ribosomes or 40 S ribosomal subunits, in combination with mouse 60 S subunits, was markedly inhibited by ethionine treatment in a complete cell-free system containing partially purified initiation factors of rabbit reticulocytes and the rat liver pH 5 fraction. (2) The polysome formation of 80 S ribosomes in the complete system described above was inhibited by ethionine treatment. Similar inhibitions by ethionine treatment were observed in the case of incubation of 40 S subunits with reticulocyte lysate, although the polysome formation was rather low even in the case of control 40 S subunits. (3) The pattern of CsCl isopycnic centrifugation of rat liver native 40 S subunits uniformly labeled with [14C]- or [3H]orotic acid showed that the content of non-ribosomal proteins of native 40 S subunits was decreased by ethionine treatment. The analysis of proteins of native 40 subunits by SDS-polyacrylamide slab gel electrophoresis revealed that eIF-3 subunits and two unidentified protein fractions of molecular weight of 2.3.10(4) and 2.1.10(4) were decreased in ethionine-treated rate liver. (4) 40 S subunits from ethionine-treated or control rat livers were labeled with N-[3H]ethylmaleimide or N-[14C]ethylmaleimide, and the 3H to 14C ratios of individual 40 S proteins on two-dimensional polyacrylamide gel electrophoresis were measured. The results suggested that the conformation of rat liver 40 S subunits was changed by ethionine treatment. (5) These results may indicate that ethionine treatment decreases the activity of rat liver 40 S subunits for the interaction with initiation factors, especially eIF-3, as the results of conformational changes of 40 S subunits.
研究了用乙硫氨酸处理雌性大鼠4小时对肝脏80S核糖体和40S核糖体亚基蛋白质合成机制的抑制作用。得到了以下结果。(1) 在含有部分纯化的兔网织红细胞起始因子和大鼠肝脏pH 5组分的完全无细胞体系中，用乙硫氨酸处理可显著抑制80S核糖体或40S核糖体亚基与小鼠60S亚基结合时对珠蛋白mRNA的翻译。(2) 上述完全体系中80S核糖体的多聚核糖体形成受到乙硫氨酸处理的抑制。在用网织红细胞裂解物孵育40S亚基的情况下，也观察到乙硫氨酸处理的类似抑制作用，尽管即使在对照40S亚基的情况下多聚核糖体形成也相当低。(3) 用[14C]-或[3H]乳清酸均匀标记的大鼠肝脏天然40S亚基的CsCl等密度离心图谱显示，乙硫氨酸处理可使天然40S亚基的非核糖体蛋白含量降低。通过SDS-聚丙烯酰胺平板凝胶电泳对天然40S亚基的蛋白质进行分析表明，在经乙硫氨酸处理的大鼠肝脏中，eIF-3亚基以及分子量分别为2.3×10(4)和2.1×10(4)的两个未鉴定蛋白组分减少。(4) 用N-[3H]乙基马来酰亚胺或N-[14C]乙基马来酰亚胺标记经乙硫氨酸处理或对照大鼠肝脏的40S亚基，并在二维聚丙烯酰胺凝胶电泳上测量各个40S蛋白的3H与14C比值。结果表明，乙硫氨酸处理可改变大鼠肝脏40S亚基的构象。(5) 这些结果可能表明，作为40S亚基构象变化的结果，乙硫氨酸处理降低了大鼠肝脏40S亚基与起始因子尤其是eIF-3相互作用的活性。
