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采用寡聚核苷酸指导的体外基因定点突变方法，将重组人肿瘤坏死因子第２０位脯氨酸变为精氨酸。突变体在大肠杆菌中的表达量是ＴＮＦＰｒｏ２０的６０％。表达产物经一系列离子交换层析分离纯化以后，突变体的比活是ＴＮＦＰｒｏ２０的千分之一。突变体和ＴＮＦＰｒｏ２０在非变性非还原的条件下进行聚丙烯酰胺凝胶电泳，两者显示相同的区带，这表明突变体也可以形成三聚体活性形式。ＴＮＦＰｒｏ２０和突变体的荧光光谱分析揭示，Ａｒｇ２０的引入改变了Ｐｒｏ２０附近的局部构象。
Using oligonucleotide-directed in vitro gene-site mutagenesis, the proline at position 20 of recombinant human tumor necrosis factor was changed to arginine. Mutants expressed in E. coli at 60% of TNFPro20. After the expressed product was isolated and purified by a series of ion exchange chromatography, the specific activity of the mutant was one thousandth of TNFPro20. Mutants and TNFPro20 were subjected to polyacrylamide gel electrophoresis under non-denaturing non-reducing conditions, both showing the same band, indicating that the mutants can also form the trimeric active form. Fluorescence spectroscopy analysis of TNFPro20 and the mutants revealed that the introduction of Arg20 altered the local conformation in the vicinity of Pro20.