Singh A, Lugovoy J M, Kohr W J, Perry L J
Nucleic Acids Res. 1984 Dec 11;12(23):8927-38. doi: 10.1093/nar/12.23.8927.
A gene fusion consisting of 960 base pairs of 5'-flanking region of the yeast MF alpha 1 gene, 257 base pairs coding for alpha-factor prepro sequence, and a modified human IFN-alpha 1 gene was constructed. MAT alpha cells containing the chimeric gene synthesized and secreted active IFN-alpha 1 into the growth medium. The secreted interferon molecules contained the last 4 amino acids of alpha-factor prepro sequence and the amino acids encoded by the DNA modifications introduced at the beginning of IFN-alpha 1 gene. DNA sequences coding for these amino acids were removed by oligonucleotide-directed in vitro mutagenesis. Yeast cells transformed with expression plasmids containing the altered junction synthesized and secreted human IFN-alpha 1 with the natural NH2-terminus.
构建了一种基因融合体，它由酵母MFα1基因5′侧翼区的960个碱基对、编码α因子前原序列的257个碱基对以及一个修饰的人IFN-α1基因组成。含有嵌合基因的MATα细胞合成并将活性IFN-α1分泌到生长培养基中。分泌的干扰素分子包含α因子前原序列的最后4个氨基酸以及在IFN-α1基因起始处引入的DNA修饰所编码的氨基酸。通过寡核苷酸定向体外诱变去除了编码这些氨基酸的DNA序列。用含有改变的连接区的表达质粒转化的酵母细胞合成并分泌具有天然NH2末端的人IFN-α1。
