Structure and catalytic mechanism of heterodisulfide reductase from methanogenic archaea

异二硫键还原酶是一种铁硫蛋白，能催化产甲烷硫代辅酶、辅酶M和辅酶B的异二硫键（CoM-S-S-Co B）的可逆还原。铁硫簇合物只能进行单电子转移反应。因此，在异二硫键还原酶中需要解决的中心问题是单电子供体如何进行二硫键的协同双电子还原。在此过程中可能形成自由基类型的中间体，例如通过二硫化物的初始单电子还原形成的硫代自由基。我们的工作假设是，这样的硫代自由基可能是稳定的铁硫簇在催化中心的酶。最近的调查已经确定了一个顺磁中心的杂二硫还原酶与EPR性能不常见的已知铁硫簇。该顺磁性中心的g值和氧化还原性质通过酶与其底物的孵育而特异性地改变。因此，它可能是催化循环中的中间体。这些发现是进一步调查的重要依据。该项目的目标是这种酶的活性位点的表征和催化循环的中间体的鉴定。

Heterodisulfide reductase from methanogenic archaea is an ironsulfur protein that can catalyze the reversible reduction of the heterodisulfide (CoM-S-S-CoB) of the methanogenic thiolcoenzymes, coenzyme M and coenzyme B. Iron-sulfur clusters can only perform one-electron transfer reactions. Thus the central problem that needs to be addressed in heterodisulfide reductase is how a one-electron donor can carry out the concerted twoelectron reduction of a disulfide. It is likely that radical type intermediates are formed during this process, for example a thiyl radical formed by the initial one-electron reduction of the disulfide. Our working hypothesis is that such a thiyl radical might be stabilized by an iron-sulfur cluster in the catalytic centre of the enzyme. Recent investigations have identified a paramagnetic centre in heterodisulfide reductase with EPR properties not common to known iron-sulfur clusters. The g-values and the redox properties of this paramagnetic centre are specifically altered by incubation of the enzyme with its substrate. It therefore might be an intermediate in the catalytic cycle. These findings are an important basis for further investigations. The objectives of this project are the characterization of the active-site of this enzyme and the identification of the intermediates of the catalytic cycle.