The preparation of rat heme oxygenase-1 mutant to reduce the level of bilirubin |
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| Authors: | Xia ZW Shao J Li YZ Chen SN Yu SC Shen QX Wang J |
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| Affiliation: | Department of Pediatrics, Rui Jin Hospital, Shanghai Second Medical University, Shanghai 200025, China;Department of Pediatrics, Rui Jin Hospital, Shanghai Second Medical University, Shanghai 200025, China;Department of Pediatrics, Rui Jin Hospital, Shanghai Second Medical University, Shanghai 200025, China;Department of Pediatrics, Rui Jin Hospital, Shanghai Second Medical University, Shanghai 200025, China;Department of Pediatrics, Rui Jin Hospital, Shanghai Second Medical University, Shanghai 200025, China;Shanghai Institute of Planned Parenthood Research, Shanghai 200032, China;Shanghai Institute of Planned Parenthood Research, Shanghai 200032, China |
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| Abstract: | OBJECTIVE: To prepare rat heme oxygenase-1 (HO-1) mutants and to determine the activity and inhibition of this mutated enzyme. METHODS: 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. RESULTS: 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%. CONCLUSION: The His25Ala mutant reduced the formation of bilirubin, suggesting that the mutant could completely bind the heme with native enzyme. |
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| Keywords: | heme oxygenase mutant hyperbilirubinemia |
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