Abstract

Chlorite dismutase is a unique heme enzyme that catalyzes the conversion of chlorite to chloride and molecular oxygen. The enzyme is highly specific for chlorite but has been known to bind several anionic and neutral ligands to the heme iron. In a pH study, the enzyme changed color from red to green in acetate buffer pH 5.0. The cause of this color change was uncovered using UV–visible and EPR spectroscopy. Chlorite dismutase in the presence of acetate showed a change of the UV–visible spectrum: a redshift and hyperchromicity of the Soret band from 391 to 404 nm and a blueshift of the charge transfer band CT1 from 647 to 626 nm. Equilibrium binding titrations with acetate resulted in a dissociation constant of circa 20 mM at pH 5.0 and 5.8. EPR spectroscopy showed that the acetate bound form of the enzyme remained high spin S = 5/2, however with an apparent change of the rhombicity and line broadening of the spectrum. Mutagenesis of the proximal arginine Arg183 to alanine resulted in the loss of the ability to bind acetate. Acetate was discovered as a novel ligand to chlorite dismutase, with evidence of direct binding to the heme iron. The green color is caused by a blueshift of the CT1 band that is characteristic of the high spin ferric state of the enzyme. Any weak field ligand that binds directly to the heme center may show the red to green color change, as was indeed the case for fluoride.

Graphic abstract

中文翻译：

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一个交通信号灯酶：乙酸酯结合可逆地将亚氯酸盐歧化酶从红色变为绿色的血红素蛋白。

摘要

亚氯酸盐歧化酶是一种独特的血红素酶，可催化亚氯酸盐转化为氯离子和分子氧。该酶对亚氯酸盐具有高度特异性，但已知可将多种阴离子和中性配体与血红素铁结合。在pH值研究中，在pH 5.0的醋酸盐缓冲液中，酶的颜色从红色变为绿色。使用紫外可见光谱和EPR光谱法可以发现这种颜色变化的原因。在乙酸盐存在下的亚氯酸盐歧化酶显示出紫外可见光谱的变化：Soret谱带从391 nm到404 nm发生红移和增色，而电荷转移谱带CT1从647到626 nm发生蓝移。用乙酸盐进行的平衡结合滴定导致pH 5.0和5.8时的解离常数约为20 mM。EPR光谱显示该酶的乙酸盐结合形式保持高自旋S  = 5/2，但是菱形明显变化，谱线变宽。近端精氨酸Arg183突变为丙氨酸导致结合乙酸盐的能力丧失。乙酸盐被发现是亚氯酸盐歧化酶的新型配体，具有直接与血红素铁结合的证据。绿色是由CT1条带的蓝移引起的，CT1带的蓝移是酶的高自旋三价铁态的特征。直接与血红素中心结合的任何弱场配体都可能显示出红色到绿色的变化，实际上氟化物就是这种情况。

图形摘要