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Relation between the structure and the oxygen acitivation of heme oxygenase reaction

血红素加氧酶反应的结构与氧活化的关系

基本信息

批准号：
12680625
负责人：
YOSHIDA Tadashi
金额：
$ 2.37万
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：
2000
资助国家：
日本
起止时间：
2000 至 2001
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-12680625/
关键词：
HEME OXYGENASE OXYGEN ACTIVATION HEME DEGRADATION INTERMEDIARY STEP BILIVERDIN REDUCTASE BILIVERDIN BILIRUBIN 酸素活性化機構ヘムの分解 CO 細菌のヘム分解酵素 Hmu O 酵素活性化機構ピリペルジンピリルビン

项目摘要

(1)The hemin complex of HmuO, a 24-kDa soluble heme degradation enzyme in Corynebacterium diphtheriae, is coordinated axially to a neutral imidazole of a proximal histidine residue in HmuO. EPR of the NO-bound ferrous heme-HmuO mutant complexes reveals His20 as the proximal heme ligand in HmuO, and this is confirmed by resonance Raman result from the ligand free ferrous heme-H20A. When bound with exogenous imidazole, His20A mutant resumes full catalytic activity. Hence, it is the absence of the proximal His ligand, that is responsible for the inactivity of proximal His mutant.(2)We found that the mutation of R183 to E or D of rat HO-1 changes the a-regioselectivity of the HO catalysis. The result shows the importance of the hydrogen bonding interaction between the arginine at position 183 and the carboxylates of the heme propionate group, as well as steric effect of the distal helix, for the a-regioselectivity.(3)Wild-type enzyme of rat HO-1 degrades heme to verdoheme with hydrogen peroxide. However, D140A heme complex forms compound II with hydrogen oeroxide, and no heme degradation occurs. D140E mutant degrades heme normally, but D140N shows reactivity similar to that of D140A. These results indicate that the carboxylate at position 140 is essential to activate the iron-bound oxygen.(4)We determined the crystal structure of rat biliverdine reductase. The structure contains two domains: an N-terminal domain characteristic of a nucleotide binding fold and a C-terminal domain. We proposed modes of binding for NADPH and biliverdin.

（1）HmuO（白喉棒状杆菌中的一种24-kDa可溶性血红素降解酶）的氯化血红素复合物轴向配位至HmuO中近端组氨酸残基的中性咪唑。NO结合亚铁血红素-HmuO突变体复合物的EPR揭示His20为HmuO中的近端血红素配体，并且这通过来自配体游离亚铁血红素-H20A的共振拉曼结果证实。当与外源咪唑结合时，His20A突变体恢复完全催化活性。因此，近端His配体的缺失是近端His突变体失活的原因。(2)We发现大鼠HO-1的R183突变为E或D改变了HO催化的α-区域选择性。结果表明，在位置183的精氨酸和血红素丙酸酯基团的羧酸酯之间的氢键相互作用，以及远端螺旋的空间效应，为a-区域选择性的重要性。（3）大鼠HO-1的野生型酶用过氧化氢将血红素降解为绿血红素。然而，D140A血红素复合物与过氧化氢形成化合物II，并且没有血红素降解发生。D140E突变体能正常降解血红素，而D140N则表现出与D140A相似的活性。这些结果表明，在位置140的羧酸是必不可少的激活铁结合氧。(4)We测定了大鼠胆绿素还原酶的晶体结构。该结构包含两个结构域：核苷酸结合折叠的N-末端结构域和C-末端结构域。我们提出了NADPH和胆绿素的结合模式。