Reaction mechanism of the iron-sulfur flavoenzyme adenosine-5′-phosphosulfate reductase based on the structural characterization of different enzymatic states
Reaction mechanism of the iron-sulfur flavoenzyme adenosine-5′-phosphosulfate reductase based on the structural characterization of different enzymatic states
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DOI：
10.1021/bi0521689
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发表时间：
2006-03-07
期刊：
影响因子：
2.9
通讯作者：
Ermler, U
中科院分区：
文献类型：
--
作者：
Schiffer, A;Fritz, G;Ermler, U
铁硫黄素酶腺苷-5'-磷酸硫酸盐 (APS) 还原酶通过将 APS 可逆地转化为亚硫酸盐和 AMP 来催化整体硫循环的关键反应。最近分别以 1.6 和 2.5 埃的分辨率阐明了还原态 (FADred) 和亚硫酸盐加合物态 (FAD-亚硫酸盐-AMP) 的古生球菌异化酶的结构。在这里，我们展示了陷入四种不同催化相关状态的酶的新结构特征，为我们提供了其反应循环的详细图片。在氧化态 (FAD(ox)) 下，FAD 辅因子的异咯嗪部分表现出与还原酶结构中观察到的类似的弯曲构象。在 APS 结合状态 (FAD(ox)-APS) 下，底物 APS 嵌入 17 A 长的底物通道中，异咯嗪环被推向通道底部，从而产生压缩的酶-底物复合物。由残基 ArgA317 和 LeuA278 形成的用于固定腺嘌呤环的钳子和弯曲的 APS 构象似乎是将 APS 保持在应变构象的关键因素。在 APS 对还原 FAD 的攻击过程中，这种能量丰富的状态被放松。在 FAD-亚硫酸盐状态的结构中观察到松弛的 FAD-亚硫酸盐加合物。最后，可以表征 FAD-亚硫酸盐-AMP1 状态，其中 AMP 在亚硫酸盐加合物的范德华距离内。该结构记录了蛋白质基质如何稳定相邻的负电荷，这对于逆反应中 AMP 和亚硫酸盐形成 APS 至关重要。
The iron-sulfur flavoenzyme adenosine-5'-phosphosulfate (APS) reductase catalyzes a key reaction of the global sulfur cycle by reversibly transforming APS to sulfite and AMP. The structures of the dissimilatory enzyme from Archaeoglobus fulgidus in the reduced state (FADred) and in the sulfite adduct state (FAD-sulfite-AMP) have been recently elucidated at 1.6 and 2.5 angstrom resolution, respectively. Here we present new structural features of the enzyme trapped in four different catalytically relevant states that provide us with a detailed picture of its reaction cycle. In the oxidized state (FAD(ox)), the isoalloxazine moiety of the FAD cofactor exhibits a similarly bent conformation as observed in the structure of the reduced enzyme. In the APS-bound state (FAD(ox)-APS), the substrate APS is embedded into a 17 A long substrate channel in such a way that the isoalloxazine ring is pushed toward the channel bottom, thereby producing a compressed enzyme-substrate complex. A clamp formed by residues ArgA317 and LeuA278 to fix the adenine ring and the curved APS conformation appear to be key factors to hold APS in a strained conformation. This energy-rich state is relaxed during the attack of APS on the reduced FAD. A relaxed FAD-sulfite adduct is observed in the structure of the FAD-sulfite state. Finally, a FAD-sulfite-AMP1 state with AMP within van der Waals distance of the sulfite adduct could be characterized. This structure documents how adjacent negative charges are stabilized by the protein matrix which is crucial for forming APS from AMP and sulfite in the reverse reaction.