Xia Z, Shao J, Shen Q, Wang J, Li Y, Chen S, Yu S
Department of Pediatrics, Rui Jin Hospital, Shanghai Second Medical University, Shanghai 200025, China.
Chin Med J (Engl). 2001 Apr;114(4):348-51.
To prepare rat heme oxygenase-1 (HO-1) mutants and to determine the activity and inhibition of this mutated enzyme.
pcDNA3HO1 containing truncated native rat HO-1 cDNA and pcDNA3HO1 delta 25 carrying mutated rat HO-1 cDNA (His25Ala) were constructed, respectively. COS-1 cells transfected with pcDNA3HO1 and pcDNA3HO1 delta 25 were collected and their activities were analyzed.
Native rat HO-1 was highly expressed in transfected cells and its activity was 13,688-15,600 U/mg protein per hour. However, the enzyme activity of mutated HO-1 declined and the value was 1948-2160 U/mg protein per hour. When an equal amount of mutant was added to the enzyme reaction system, the level of bilirubin decreased by 42%.
The His25Ala mutant reduced the formation of bilirubin, suggesting that the mutant could completely bind the heme with native enzyme.
制备大鼠血红素加氧酶-1（HO-1）突变体，并测定该突变酶的活性及抑制作用。
分别构建含截短的天然大鼠HO-1 cDNA的pcDNA3HO1和携带突变大鼠HO-1 cDNA（His25Ala）的pcDNA3HO1 delta 25。收集用pcDNA3HO1和pcDNA3HO1 delta 25转染的COS-1细胞，并分析其活性。
天然大鼠HO-1在转染细胞中高表达，其活性为每小时13,688 - 15,600 U/mg蛋白。然而，突变型HO-1的酶活性下降，值为每小时1948 - 2160 U/mg蛋白。当向酶反应体系中加入等量突变体时，胆红素水平下降了42%。
His25Ala突变体减少了胆红素的形成，表明该突变体可与天然酶完全结合血红素。
