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目的构建一种新型人肿瘤坏死因子（ＴＮＦα）分子，测定其蛋白表达和生物学活性。方法在详细分析ＴＮＦα结构及其结构与功能关系的基础上，设计并人工合成了一对ＴＮＦα结构基因点突变引物。应用ＰＣＲ分子克隆技术构建了一种新型ＴＮＦα分子的编码基因，将该编码基因插入表达质粒，转化大肠杆菌，通过温度诱导获得了表达蛋白，对表达产物进行免疫学检测和生物活性检测。结果新构建的突变体ＴＮＦαΝ１与ＴＮＦαＥＬＩＳＡ试剂盒有免疫学反应，其含量为０．５μｇ／ｍｌ；在体外有生物学活性，为１０７ＢＵ／ｍｌ，比活为２．１×１０５ＢＵ／ｍｇ；理化性质与天然原型ＴＮＦα有所不同。结论得到了一种具有生物学活性的新型ＴＮＦα分子。
Objective To construct a new type of human tumor necrosis factor (TNF α) molecule and determine its protein expression and biological activity. Methods Based on a detailed analysis of the relationship between TNFα structure and its structure and function, a pair of TNFα structural gene point mutation primers were designed and synthesized. A novel TNFα gene encoding gene was constructed by PCR molecular cloning technique. The gene was inserted into the expression plasmid and transformed into E. coli. The expression protein was obtained by temperature induction, and the expression product was detected by immunology and biological activity. Results The newly constructed mutant TNF TNFα TNF α ELISA ELISA kit with immunological reactions, its content 0.5μg / ml; in vitro biological activity of 107BU / ml, the specific activity of 2.1 × 105BU / mg; physical and chemical properties and natural prototype TNF @ different. Conclusions A novel TNFα molecule with biological activity was obtained.