Structure and catalytic mechanism of heterodisulfide reductase from methanogenic archaea

来自产甲烷古菌的异二硫还原酶是一种铁硫蛋白，可以催化产甲烷硫醇酶、辅酶M和辅酶b的异二硫（CoM-S-S-CoB）的可逆还原。因此，在杂二硫还原酶中需要解决的中心问题是一个单电子供体如何对二硫进行协调的双电子还原。在此过程中很可能形成自由基型中间体，例如由二硫化合物初始的单电子还原形成的巯基自由基。我们的工作假设是，这种硫基自由基可能被酶催化中心的铁硫簇稳定下来。最近的研究已经确定了一个顺磁中心在异二硫还原酶与EPR性质不常见的已知铁硫簇。这种顺磁中心的g值和氧化还原性质在酶与其底物的孵育过程中被特异性地改变。因此它可能是催化循环中的中间体。这些发现是进一步研究的重要基础。该项目的目的是表征该酶的活性位点和鉴定催化循环的中间体。

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.