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

血红素加氧酶通过三个顺序的单加氧酶反应催化氯化血红素的区域特异性氧化生成双折叠IXα，同时释放CO和铁。(1)用电子顺磁共振研究了血红素转化为α-中羟基血红素的反应。Endor和光学吸收光谱。这些研究结果表明，氢过氧铁-HO确实是活性物种，通过氢过氧基部分的远端羟基在血红素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)利用截短的大鼠HO-1和化学合成的四种异构体，研究了从中羟基氯化血红素到马鞭铁血红素和马鞭铁血红素到胆绿素的两个反应步骤的立体选择性。HO-1将中羟基氯化高铁血红素的四种异构体全部转化为相应的…更多的马鞭草。相反，只有Verdoheme IXα被转化为相应的胆绿素IXα。我们得出结论，对于α-异构体底物来说，第三步是立体选择性的，而不是第二步。(3)蓝藻聚胞藻高效表达系统。利用人工合成的基因构建了PCC6803HO基因ho-1。首次获得了高水平表达和高度纯化的可溶性蛋白。光谱特征和分解代谢活性强烈表明，尽管聚球藻HO-1的一级结构非常保守，但其血红素口袋结构与大鼠HO的不同。(4)白喉棒杆菌HO的铁和亚铁血红素络合物的晶体结构分别被细化到1.4和1.5A。血红素口袋结构负责通过防止过早的水解性O-O键断裂来稳定铁的过氧氢活性中间体。较少

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