Hough C J, Chaiken I M
Nucleic Acids Res. 1984 May 25;12(10):4397-410. doi: 10.1093/nar/12.10.4397.
The coding activity of bovine hypothalamic poly A+ mRNA for neurophysin I and II immunoreactive proteins was characterized with respect to size and 5' cap. The mRNA was fractionated by methylmercuric hydroxide agarose gel electrophoresis and subsequently translated in vitro in rabbit reticulocyte lysates. Alternatively, mRNA was fractionated by gel exclusion HPLC and translated in wheat germ extracts. Immunoprecipitated translation products were analyzed by gel exclusion HPLC. Neurophysin-immunospecific protein of approximately 17,000 daltons, the size expected for the neuropeptide hormone-neurophysin precursors, was encoded by mRNA species of two size classes. The smaller class of mRNA's was of the size expected from the size of the precursor proteins. The larger class was 5-10 times larger. The low K+ concentration optimum for translation of unfractionated mRNA encoding neurophysin I immunoreactive proteins and the inability of a cap analogue to inhibit this translation suggest that mRNA species encoding neurophysin I-immunoreactive translation products are incompletely capped. By contrast, the mRNA encoding neurophysin II-immunoreactive products appear to contain a normal cap structure.
对牛下丘脑多聚腺苷酸加尾mRNA编码神经垂体素I和II免疫反应性蛋白的编码活性,在大小和5'帽结构方面进行了表征。该mRNA通过氢氧化甲基汞琼脂糖凝胶电泳进行分级分离,随后在兔网织红细胞裂解物中进行体外翻译。或者,mRNA通过凝胶排阻高效液相色谱法进行分级分离,并在小麦胚芽提取物中进行翻译。通过凝胶排阻高效液相色谱法分析免疫沉淀的翻译产物。约17,000道尔顿的神经垂体素免疫特异性蛋白,即神经肽激素 - 神经垂体素前体预期的大小,由两种大小类别的mRNA编码。较小类别的mRNA大小与前体蛋白大小预期相符。较大类别的mRNA比预期大5至10倍。编码神经垂体素I免疫反应性蛋白的未分级mRNA翻译的最佳钾离子浓度较低,且帽类似物无法抑制这种翻译,这表明编码神经垂体素I免疫反应性翻译产物的mRNA种类帽结构不完全。相比之下,编码神经垂体素II免疫反应性产物的mRNA似乎含有正常的帽结构。