Influence of the Environment on the [4Fe-4S]2+ to [2Fe-2S]2+ Cluster Switch in the Transcriptional Regulator FNR

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• 作者：Jason C. Crack ; Alisa A. Gaskell ; Jeffrey Green ; Myles R. Cheesman ; Nick E. Le Brun ; Andrew J. Thomson
• 刊名：Journal of the American Chemical Society
• 出版年：2008
• 出版时间：February 6, 2008
• 年：2008
• 卷：130
• 期：5
• 页码：1749 - 1758
• 全文大小：614K
• 年卷期：v.130,no.5(February 6, 2008)
• ISSN：1520-5126

文摘

In Escherichia coli, the switch between aerobic and anaerobic metabolism is primarily controlledby the fumarate and nitrate reduction transcriptional regulator FNR. In the absence of O₂, FNR binds a[4Fe-4S]²⁺ cluster, generating a transcriptionally active dimeric form. Exposure to O₂ results in theconversion of the cluster to a [2Fe-2S]²⁺ form, leading to dissociation of the protein into transcriptionallyinactive monomers. The [4Fe-4S]²⁺ to [2Fe-2S]²⁺ cluster conversion proceeds in two steps. Step 1 involvesthe one-electron oxidation of the cluster, resulting in the release of Fe²⁺, generating a [3Fe-4S]¹⁺ clusterintermediate, and a superoxide ion. In step 2, the cluster intermediate spontaneously rearranges to formthe [2Fe-2S]²⁺ cluster, with the release of a Fe³⁺ ion and two sulfide ions. Here, we demonstrate that, inboth native and reconstituted [4Fe-4S] FNR, the reaction environment and, in particular, the presence ofFe²⁺ and/or Fe³⁺ chelators can influence significantly the cluster conversion reaction. We demonstratethat while the rate of step 1 is largely insensitive to chelators, that of step 2 is significantly enhanced byboth Fe²⁺ and Fe³⁺ chelators. We show that, for reactions in Fe³⁺-coordinating phosphate buffer, step 2 isenhanced to the extent that step 1 becomes the rate determining step and the [3Fe-4S]¹⁺ intermediate isno longer detectable. Furthermore, Fe³⁺ released during this step is susceptible to reduction in the presenceof Fe²⁺ chelators. This work, which may have significance for the in vivo FNR cluster conversion reactionin the cell cytoplasm, provides an explanation for apparently contradictory results reported from differentlaboratories.