虫荧光素酶N末端缺失

N terminal deletion of firefly luciferase

目的是通过虫荧光素酶Ｎ末端氨基酸的缺失，研究缺失对酶活性的影响。采用聚合酶链式反应的方法，构建虫荧光素酶Ｎ－末端缺失７，８，９个氨基酸残基的突变体，导入大肠杆菌ＤＨ５α菌株中直接得到表达，再分离纯化粗酶，并检测酶活。结果表明，Ｎ末端缺失７个氨基酸的突变体几乎没有活性（小于天然活性的０．５％），Ｎ末端缺失８个以上氨基酸残基的突变体则完全丧失活性。由于Ｎ末端缺失６个氨基酸的突变体保持了７７％的酶活性，因而萤火虫荧光素酶Ｎ末端第７个氨基酸与酶的催化活性密切相关。

The article researches the affect deletion of N terminal amino acids of firefly luciferase on activity of the enzyme. The gene mutants with N terminal deletion of the first 7, 8 and 9 amino acids of firefly luciferase were synthesized by polymerase chain reaction (PCR) and conducted into E.coli strain DH5α. The expressed enzyme extracts were purified and the enzymatic activities were examined qualitatively. Final results showed that the mutant with N terminal deletion of the first 7 amino acids maintained trace enzymatic activity (less than 0.5%), while the mutant losing the first more than 8 amino acids was robbed of the enzymatic function completely. Since the mutant lossing the first 6 amino acids from the N terminus of firefly luciferase keeps 77% of natural luciferase activity, the 7th N terminal amino acid is of great importance to the normal enzymatic function.