Catalytic enhancement of the heme-based oxygen-sensing phosphodiesterase EcDOS by hydrogen sulfide is caused by changes in heme coordination structure.
Catalytic enhancement of the heme-based oxygen-sensing phosphodiesterase EcDOS by hydrogen sulfide is caused by changes in heme coordination structure.
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硫化氢对血红素氧敏磷酸二酯酶 EcDOS 的催化增强是由血红素配位结构的变化引起的。
DOI：
10.1007/s10534-015-9847-7
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发表时间：
2015
期刊：
影响因子：
3.5
通讯作者：
Shimizu Toru
中科院分区：
文献类型：
--
作者：
Yan Fang;Fojtikova Veronika;Man Petr;Stranava Martin;Martinkova Marketa;Du Yongming;Huang Dongyang;Shimizu Toru
EcDOS是一种以血红素为基础的o2感应磷酸二酯酶，其中o2与血红素铁复合物在n端结合，大大增强了c端结构域对环二gmp的催化作用。在这里，我们发现硫化氢增强了全长cdos的催化活性，可能是由于反应过程中生成的6配位血红素Fe(III) -SH -和Fe(II) - o2配合物的混合物。Met95是血红素Fe（II）复合物的轴向配体，其丙氨酸取代将血红素Fe（III）复合物转化为血红素Fe(III) -SH -复合物，但Na2S的加入并没有进一步将其还原为Met95Ala突变体的血红素Fe（II）复合物，也没有观察到随后形成血红素Fe(II) - o2复合物。相比之下，Met95His突变体在同样的处理下形成了稳定的血红素Fe(II) - o2复合物。Arg97Glu突变体在血红素Fe(II) - O2复合物中与O2相互作用的氨基酸上含有谷氨酸取代，在Na2S作用下形成稳定的血红素Fe（II）复合物，但该复合物不能结合O2。有趣的是，在Arg97Ile突变体中，Na2S的加入促进了verdoheme（含氧修饰的原卟啉IX）的形成。Na2S对Met95突变体和野生型的催化增强作用相似，但对Arg97突变体的催化增强作用明显较弱。因此，本研究首次分离到光谱分离的稳定的血红素Fe(III) -SH−、血红素Fe（II）和血红素Fe(II) - o2与Na2S全长的dos配合物，并证实了外配体结合的6配位血红素Fe(III) -SH−或血红素Fe(II) - o2配合物对Na2S诱导的ecdos催化增强起关键作用。
EcDOS is a heme-based O2-sensing phosphodiesterase in which O2binding to the heme iron complex in the N-terminal domain substantially enhances catalysis toward cyclic-di-GMP, which occurs in the C-terminal domain. Here, we found that hydrogen sulfide enhances the catalytic activity of full-lengthEcDOS, possibly owing to the admixture of 6-coordinated heme Fe(III)–SH−and Fe(II)–O2complexes generated during the reaction. Alanine substitution at Met95, the axial ligand for the heme Fe(II) complex, converted the heme Fe(III) complex into the heme Fe(III)–SH−complex, but the addition of Na2S did not further reduce it to the heme Fe(II) complex of the Met95Ala mutant, and no subsequent formation of the heme Fe(II)–O2complex was observed. In contrast, a Met95His mutant formed a stable heme Fe(II)–O2complex in response to the same treatment. An Arg97Glu mutant, containing a glutamate substitution at the amino acid that interacts with O2in the heme Fe(II)–O2complex, formed a stable heme Fe(II) complex in response to Na2S, but this complex failed to bind O2. Interestingly, the addition of Na2S promoted formation of verdoheme (oxygen-incorporated, modified protoporphyrin IX) in an Arg97Ile mutant. Catalytic enhancement by Na2S was similar for Met95 mutants and the wild type, but significantly lower for the Arg97 mutants. Thus, this study shows the first isolation of spectrometrically separated, stable heme Fe(III)–SH−, heme Fe(II) and heme Fe(II)–O2complexes of full-lengthEcDOS with Na2S, and confirms that external-ligand–bound, 6-coordinated heme Fe(III)–SH−or heme Fe(II)–O2complexes critically contribute to the Na2S-induced catalytic enhancement ofEcDOS.