Iron(II)- and 2-(oxo)-glutarate-dependent (Fe/2OG) oxygenases catalyze a diverse array of oxidation reactions via a common iron(IV)-oxo (ferryl) intermediate. Although the intermediate has been characterized spectroscopically, its short lifetime has precluded crystallograhic characterization. In solution, the ferryl was first observed directly in the archetypal Fe/2OG hydroxylase, taurine:2OG dioxygenase (TauD). Here, we substitute the iron cofactor of TauD with the stable vanadium(IV)-oxo (vanadyl) ion to obtain crystal structures mimicking the key ferryl complex. Intriguingly, whereas the structure of the TauD·(V^IV-oxo)·succinate·taurine complex exhibits the expected orientation of the V≡O bond—trans to the His255 ligand and toward the C–H bond to be cleaved, in what has been termed the in-line configuration—the TauD·(V^IV-oxo) binary complex is best modeled with its oxo ligand trans to Asp101. This off-line-like configuration is similar to one recently posited as a means to avoid hydroxylation in Fe/2OG enzymes that direct other outcomes, though neither has been visualized in an Fe/2OG structure to date. Whereas an off-line (trans to the proximal His) or off-line-like (trans to the carboxylate ligand) ferryl is unlikely to be important in the hydroxylation reaction of TauD, the observation that the ferryl may deviate from an in-line orientation in the absence of the primary substrate may explain the enzyme’s mysterious self-hydroxylation behavior, should the oxo ligand lie trans to His99. This finding reinforces the potential for analogous functional off-line oxo configurations in halogenases, desaturases, and/or cyclases.

中文翻译：

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原型铁（II）-和2-（Oxo）-戊二酸依赖性双加氧酶TauD中的轮渡模拟物的结构。

依赖于铁（II）-和2-（氧代）-谷氨酸的（Fe / 2OG）加氧酶可通过常见的铁（IV）-氧代（轮式）中间体催化各种氧化反应。尽管该中间体已通过光谱表征，但其使用寿命短，无法进行结晶学表征。在溶液中，首先在原型Fe / 2OG羟化酶，牛磺酸：2OG双加氧酶（TauD）中直接观察到小轮。在这里，我们用稳定的钒（IV）-氧代（钒基）离子代替了TauD的铁辅因子，从而获得了模拟关键小轮配合物的晶体结构。有趣的是，而TAUD的结构·（V ^IV -氧）·琥珀酸酯·牛磺酸复合物表现出V≡O成键的预期定向反称为直链构型的TauD·（V ^IV -oxo ）二元配合物最好是通过将其oxo配体反式转化为Asp101来形成的。这种离线样的构型类似于最近被定位为避免指导其他结果的Fe / 2OG酶中羟基化的一种构型，尽管迄今为止在Fe / 2OG结构中都没有观察到这种构型。脱机（反接至近端His）或脱机式（反接对于羧酸盐配体），在TauD的羟基化反应中，亚铁不太可能重要。如果没有主要底物，则亚铁可能偏离在线方向，这一观察结果可以解释该酶的神秘自羟基化行为。氧代配体位于反义到His99。这一发现增强了卤化酶，去饱和酶和/或环化酶中类似的功能性离线羰基合成构型的潜力。