Mechanism of heme degradation by heme oxygenase and the interaction of heme

血红素加氧酶降解血红素的机制及血红素的相互作用

• 批准号: 14580641
• 负责人: YOSHIDA Tadashi
• 金额: $ 2.62万
• 依托单位: Yamagata University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2002
• 资助国家: 日本
• 起止时间: 2002 至 2003
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-14580641/
• 关键词: HEME OXYGENASE; OXYGEN ACTIVATION; HYDROPEROXO SPECIES; meso-HYDROXYHEME; VERDOHEME; BILIVERDIN; CYANOBACTERIUM; Corynebacterium diphtheriae; 酸素活性化機構; ヘムの分解; CO; ビリルビン; ビリベルジン還元酵素

Heme oxygenase catalyzes the regiospecific oxidation of hemin to biliverding IXα with concomitant liberation of CO and iron by three sequential mono-oxygenase reaction. (1) We studied the conversion of heme to α-meso-hydroxyheme by EPR. ENDOR, and optical absorption spectroscopy. The results obtained by these studies demonstrate that hydroperoxo ferric-HO is indeed the reactive species, directly forming the α-meso-hydroxyheme by attack of the distal OH of hydroperoxo moiety at the heme α-carbon. D140A mutant failed to convert heme to α-meso-hydroxyheme, confirming our previous finding that the H-bonding network within the distal pocket of HO is disrupted. (2) We investigated the stereoselectivity of each of the two reaction steps from meso-hydroxyhemin to verdoheme and versoheme to biliverdin, by using a truncated form of rat HO-1 and the chemically synthesized four isomers of meso-hydroxythemin and verdoheme. HO-1 converted all four isomers of meso-hydroxyhemin to the corresponding is … More omers of verdoheme. In contrast, only verdoheme IXα was converted to the corresponding biliverdin IXα. We conclude that the third step, but not the second, is stereoselective for the α-isomer substrate. (3) An efficient bacterial expression system of cyanobacterium Synechocystis sp. PCC 6803 HO gene, ho-1, has been constructed, using a synthetic gene. A soluble protein was expressed at high levels and was highly purified, for the first time. The spectroscopic characters as well as the catabolic activities strongly suggest that, in spite of very high conservation of the primary structure, the heme pocket structure of Synechocystis HO-1 is different from that of rat HO. (4) Crystal structure of the ferric and ferrous heme complexes of HemO, a 24-kDA HO of Corynebacterium diphtheriae, have been refined to 1.4 and 1.5 A resolustion, respectively. The heme pocket architecture is responsible for stabilization of the ferric hydroperoxo-active intermediated by preventing premature hydrolytic O-O bond cleavage. Less

血红素加氧酶通过三个连续的单加氧酶反应催化血红素的区域特异性氧化生成IXα，同时释放CO和铁。(1)利用EPR研究了血红素向α-中羟基血红素的转化。和光学吸收光谱。这些研究结果表明，氢过氧铁- ho确实是反应物质，通过氢过氧部分的远端OH在血红素α-碳上的攻击直接形成α-中羟基血红素。D140A突变体未能将血红素转化为α-中羟基血红素，证实了我们之前的发现，即HO远端口袋内的氢键网络被破坏。(2)利用大鼠HO-1的截断形式和化学合成的中羟基血红素和蛇毒血红素的四种异构体，研究了中羟基血红素到蛇毒血红素和蛇毒血红素到胆绿素两个反应步骤的立体选择性。HO-1将中羟血红素的所有四种异构体转化为相应的四种羟血红素异构体。相反，只有胆绿素IXα转化为胆绿素IXα。我们得出结论，第三步，而不是第二步，对α-异构体底物具有立体选择性。(3)利用人工合成的基因构建了蓝细菌聚囊藻PCC 6803 HO基因HO -1的高效表达体系。首次高水平表达了可溶性蛋白，并进行了高纯度纯化。光谱学特征和分解代谢活性强烈表明，尽管HO-1的血红素袋结构高度保守，但它与大鼠HO的血红素袋结构不同。(4)白喉棒状杆菌的24 kda HO - HemO的铁血红素和铁血红素配合物的晶体结构分别被细化到1.4和1.5 a分辨率。血红素口袋结构通过防止过早水解O-O键断裂来稳定铁的氢过氧活性中间体。少

项目成果

Denisov, IG.: "Cryogenic absorption spectra of hydroperoxo-ferric heme oxygenase"FEBS Letters. 532. 203-206 (2002)

Denisov，IG.：“氢过氧铁血红素加氧酶的低温吸收光谱”FEBS 快报。

Hiroshi Fujii: "Essential amino acid residues controlling the unique regioselectivity of heal oxygenase in Psoudommes aeruginase"Journal of American Chemical Society. 126(印刷中). (2004)

Hiroshi Fujii：“控制铜绿假单胞菌氧化酶独特区域选择性的必需氨基酸残基”美国化学会杂志 126（出版中）。

Xuhong Zhang: "Stereoselectivity of each of the three steps of the heme oxygenase reaction : Hemin to meso-hydroxyhemin, to verdoheme"Biochemistry. 42. 7418-7426 (2003)

张旭红：“血红素加氧酶反应三个步骤中每一步的立体选择性：血红素到内消旋羟基血红素，到绿血红素”生物化学。

Shinji Susa: "Heme inhibits the mitochondrial import of coproporphyrinogen oxidase"BLOOD. 100・13. 4678-4679 (2002)

Shinji Susa：“血红素抑制粪卟啉原氧化酶的线粒体输入”BLOOD 100・13（2002）。

Denisov IG et al.: "Cryogenic absorption spectra of hydroperoxo-ferric heme oxygenase, the active intermediate of enzymatic heme oygenatgion."FEBS Lett. 532. 203-206 (2002)

Denisov IG 等人：“氢过氧铁血红素加氧酶的低温吸收光谱，它是酶促血红素加氧的活性中间体。”FEBS Lett。