Zitomer R S, Hall B D
J Biol Chem. 1976 Oct 25;251(20):6320-6.
An assay based upon indirect immunoprecipitation has been developed for yeast cytochrome c and apocytochrome c. The specificity of this assay was demonstrated by its ability to selectively precipitate cytochrome c from an autolysate of yeast cell proteins. Translation of the polypeptide chain of cytochrome c in a wheat germ extract programmed with yeast poly(A) RNA was demonstrated using this immunoprecipitation assay. Translation of poly(A) RNA from yeast strains carrying nonsense mutations in the cyc1 gene yielded in vitro cytochrome c polypeptides which were shorter than the wild type protein by the amount expected for polypeptide chains which had terminated at the nonsense codon. The in vivo rate of cytochrome c synthesis was shown to be 6-fold greater in derepressed cells than in glucose-repressed cells. The 6-fold difference is sufficient to account for the 6-fold higher level of cytochrome c in derepressed than in repressed cells. The level of translatable cytochrome c mRNA is at least 4 times as high in derepressed as in glucose-repressed cells, suggesting that regulation occurs at some step in the synthesis of this messenger.
已开发出一种基于间接免疫沉淀的方法来检测酵母细胞色素c和脱辅基细胞色素c。该方法的特异性通过其从酵母细胞蛋白自溶产物中选择性沉淀细胞色素c的能力得以证明。利用这种免疫沉淀方法，证实了用酵母聚腺苷酸RNA编程的小麦胚芽提取物中细胞色素c多肽链的翻译过程。来自在cyc1基因中携带无义突变的酵母菌株的聚腺苷酸RNA的翻译产生了体外细胞色素c多肽，其比野生型蛋白短，缩短的长度与在无义密码子处终止的多肽链预期的长度一致。结果表明，去阻遏细胞中细胞色素c的体内合成速率比葡萄糖阻遏细胞中的合成速率高6倍。这6倍的差异足以解释去阻遏细胞中细胞色素c水平比阻遏细胞中高6倍的现象。去阻遏细胞中可翻译的细胞色素c信使核糖核酸（mRNA）水平至少是葡萄糖阻遏细胞中的4倍，这表明调控发生在该信使合成的某个步骤。
