The preparation of rat heme oxygenase-1 mutant to reduce the level of bilirubin
Xia, Z.; Shao, J.; Shen, Q.; Wang, J.; Li, Y.; Chen, S.; Yu, S.
Chinese Medical Journal 114(4): 348-351
2001
ISSN/ISBN: 0366-6999 PMID: 11780451 Document Number: 533459
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.