Substrate-Triggered Addition of Dioxygen to the Diferrous Cofactor of Aldehyde-Deformylating Oxygenase to Form a Diferric-Peroxide Intermediate

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• 作者：Maria E. Pandelia ; Ning Li ; Hanne N酶rgaard ; Douglas M. Warui ; Lauren J. Rajakovich ; Wei-chen Chang ; Squire J. Booker ; Carsten Krebs ; J. Martin Bollinger ; Jr.
• 刊名：Journal of the American Chemical Society
• 出版年：2013
• 出版时间：October 23, 2013
• 年：2013
• 卷：135
• 期：42
• 页码：15801-15812
• 全文大小：472K
• 年卷期：v.135,no.42(October 23, 2013)
• ISSN：1520-5126

文摘

Cyanobacterial aldehyde-deformylating oxygenases (ADOs) belong to the ferritin-like diiron-carboxylate superfamily of dioxygen-activating proteins. They catalyze conversion of saturated or monounsaturated C_n fatty aldehydes to formate and the corresponding C_n鈥? alkanes or alkenes, respectively. This unusual, apparently redox-neutral transformation actually requires four electrons per turnover to reduce the O₂ cosubstrate to the oxidation state of water and incorporates one O-atom from O₂ into the formate coproduct. 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 Fe₂^III/III complex with a bound peroxide. Its M枚ssbauer spectra reveal that the intermediate possesses an antiferromagnetically (AF) coupled Fe₂^III/III center with resolved subsites. The intermediate is long-lived in the absence of a reducing system, decaying slowly (t_1/2 400 s at 5 掳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-methylphenazinium methylsulfate (^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 heterodinuclear 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 Fe₂^III/III-peroxyhemiacetal complex, which undergoes reductive O鈥揙-bond cleavage, leading to C1鈥揅2 radical fragmentation and formation of the alk(a/e)ne and formate products.