Substrate-triggered addition of dioxygen to the diferrous cofactor of aldehyde-deformylating oxygenase to form a diferric-peroxide intermediate.
Substrate-triggered addition of dioxygen to the diferrous cofactor of aldehyde-deformylating oxygenase to form a diferric-peroxide intermediate.
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DOI：
10.1021/ja405047b
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发表时间：
2013-10-23
影响因子：
15
通讯作者：
Bollinger JM Jr
中科院分区：
文献类型：
--
作者：
Pandelia ME;Li N;Nørgaard H;Warui DM;Rajakovich LJ;Chang WC;Booker SJ;Krebs C;Bollinger JM Jr
蓝藻脱羧脱甲酰加氧酶（ADOs）属于铁蛋白样二铁羧酸盐超家族的双氧激活蛋白。它们分别催化饱和或单不饱和Cn脂肪醛转化为甲酸酯和相应的Cn-1烷烃或烯烃。这种不寻常的，显然是氧化还原中性的转变实际上需要四个电子每次周转，以减少O2共底物的水的氧化态，并将一个O-原子从O2到甲酸盐的副产品。在这里，我们表明，从念珠藻（NP）与醛底物的ADO的二铁（II/II）形式的复合物与O2反应，形成一个有色的中间体与光谱特性暗示的Fe 2 III/III复合物与绑定过氧化物。其穆斯堡尔谱表明，该中间体具有反铁磁（AF）耦合的Fe 2 III/III中心与解决的子网站。中间体在没有还原系统的情况下是长寿命的，缓慢地衰变（t1/2 ~ 400 s，5 °C），以产生非常适度的甲酸盐产率（< 0.15酶当量），但与完全还原形式的1-甲氧基-5-甲基吩嗪（MeOPMS）快速反应，产生产物，尽管仅为最大理论产率的~ 50%（由于来自一种或多种非生产性途径的竞争）。这些结果代表了迄今为止最明确的证据，即ADO可以使用二铁辅因子（而不是涉及另一种过渡金属的同或异双核簇），并为涉及通过还原的O2部分攻击结合底物的羰基以形成Fe 2 III/III-过氧半缩醛络合物的机制提供支持，所述Fe 2 III/III-过氧半缩醛络合物经历还原性O-O键断裂，导致C1-C2自由基断裂并形成烷（a/e）ne和甲酸盐产物。
Cyanobacterial aldehyde-deformylating oxygenases (ADOs) belong to the ferritin-like diiron-carboxylate superfamily of dioxygen-activating proteins. They catalyze conversion of saturated or mono-unsaturated Cn fatty aldehydes to formate and the corresponding Cn-1 alkanes or alkenes, respectively. This unusual, apparently redox-neutral transformation actually requires four electrons per turnover to reduce the O2 co-substrate to the oxidation state of water and incorporates one O-atom from O2 into the formate co-product. We show here that the complex of the diiron(II/II) form of ADO from Nostoc punctiforme (Np) with an aldehyde substrate reacts with O2 to form a colored intermediate with spectroscopic properties suggestive of a Fe2III/III complex with a bound peroxide. Its Mössbauer spectra reveal that the intermediate possesses an antiferromagnetically (AF) coupled Fe2III/III center with resolved sub-sites. The intermediate is long-lived in the absence of a reducing system, decaying slowly (t1/2 ~ 400 s at 5 °C) to produce a very modest yield of formate (< 0.15 enzyme equivalents), but reacts rapidly with the fully reduced form of 1-methoxy-5-methylphenazine (MeOPMS) to yield product, albeit at only ~ 50% of the maximum theoretical yield (owing to competition from one or more unproductive pathway). The results represent the most definitive evidence to date that ADO can use a diiron cofactor (rather than a homo- or hetero-dinuclear cluster involving another transition metal) and provide support for a mechanism involving attack on the carbonyl of the bound substrate by the reduced O2 moiety to form a Fe2III/III-peroxyhemiacetal complex, which undergoes reductive O-O-bond cleavage, leading to C1–C2 radical fragmentation and formation of the alk(a/e)ne and formate products.