2005 Fiscal Year Final Research Report Summary
Roles of novel hydroperoxo-type active intermediates in the heme oxygenase catalysis
| Project/Area Number |
16370056
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Functional biochemistry
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| Research Institution | Tohoku University |
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Principal Investigator |
SAITO Masao Tohoku University, Institute of Multidisciplinary Research for Advanced Materials, Professor, 多元物質科学研究所, 教授 (70302239)
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| Co-Investigator(Kenkyū-buntansha) |
MATSUI Toshitaka Tohoku University, Institute of Multidisciplinary Research for Advanced Materials, Lecturer, 多元物質科学研究所, 講師 (90323120)
UNNO Masaki Tohoku University, Institute of Multidisciplinary Research for Advanced Materials, Research associate, 多元物質科学研究所, 助手 (10359549)
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| Project Period (FY) |
2004 – 2005
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| Keywords | heme / heme oxygenase / X-ray crystallography / reaction mechanism / oxygen activation |
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| Research Abstract |
Heme oxygenase (HO) degrades heme to biliverdin through three successive oxygenations. Specific aim of this research project is to delineate the first and third oxygenations by HO (heme hydroxylation and verdoheme ring opening) through detailed reaction analysis, crystallographic and spectroscopic characterization. Our special attention has been paid to elucidate formation and activation mechanisms of novel hydroperoxo active intermediates.
In the first oxygenation, the hydroperoxo species has been proposed as an active species. Although the hydroperoxo may transiently afford an oxoferryl porphyrin cation radical (so called compound I), we have found that compound I of HO never hydroxylates heme. We then have analyzed formation and activation process of the hydroperoxo complex in detail. Upon the cryo-reduction of oxy-heme complex, the peroxo moiety is immediately protonated even at 4 K by an adjacent water molecule which consists a hydrogen bond network in the HO active site. The water again protonates the hydroperoxo species formed to enable the self-hydroxylation of heme. The oxy-HO in crystal can be transformed by cryo-X-ray irradiation to the hydroperoxo-HO whose structure is successfully determined at 1.65 Å resolution. The unique conformation of the Fe-O-O moiety can be critical for the self-hydroxylation.
The third oxygenation, a major rate-determining step, is the least understood step in the HO catalysis. Our reaction analysis found that HO degrades verdoheme through a dual pathway using either O_2 or H_2O_2 with accumulation of distinct intermediary species. Inhibition and mutagenesis studies strongly suggest the formation of a hydroperoxo species as a key intermediate. These results including kinetic parameters of the ring-opening reaction delineate the regulation mechanism of the HO enzyme to help understand the biological functions of HO.
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