Carmichael G, Schaffhausen B S, Mandel G, Liang T J, Benjamin T L
Proc Natl Acad Sci U S A. 1984 Feb;81(3):679-83. doi: 10.1073/pnas.81.3.679.
We used an oligonucleotide to introduce an A----T transversion at nucleotide position 1178 in polyoma virus DNA. The single effect of this mutation is to substitute phenylalanine for tyrosine at residue 315 of the middle-sized tumor (mT) protein (antigen). This site was previously identified as a major phosphate acceptor in the protein kinase reaction of immunocomplexes containing mT antigen. Reconstituted polyoma virus with the transversion, Py-1178-T, produces an altered mT protein that shows about 20% of the activity of wild-type mT antigen in the immunocomplex kinase assay. This residual activity appears to be directed primarily at another tyrosine at position 322 in the mT protein. The transforming ability of Py-1178-T is drastically reduced compared to wild-type virus. The efficiency of transformation by the mutant is less than 1% of that of wild type in focus assays and less than 0.1% in soft-agar growth assays. Cells identified in focus assays with Py-1178-T are generally less transformed in their phenotype than wild-type transformed cells.
我们使用一种寡核苷酸在多瘤病毒DNA的第1178位核苷酸处引入A到T的颠换。这种突变的单一效应是在中等大小肿瘤（mT）蛋白（抗原）的第315位残基处将酪氨酸替换为苯丙氨酸。该位点先前被确定为含有mT抗原的免疫复合物蛋白激酶反应中的主要磷酸受体。携带这种颠换的重组多瘤病毒Py-1178-T产生一种改变的mT蛋白，在免疫复合物激酶测定中显示出约20%的野生型mT抗原活性。这种残余活性似乎主要针对mT蛋白中第322位的另一个酪氨酸。与野生型病毒相比，Py-1178-T的转化能力大幅降低。在焦点试验中，突变体的转化效率不到野生型的1%，在软琼脂生长试验中不到0.1%。用Py-1178-T在焦点试验中鉴定出的细胞，其表型通常比野生型转化细胞的转化程度低。
