Studies on the enzymatic chemistry involved in the inorganic sulfur metabolism and its application

本研究旨在提高我们对硫氧化菌中参与异化无机硫代谢的酶的认识。我们分离并鉴定了四硫酸水解酶（af - th）的基因，该酶是酸性氧化亚铁硫杆菌代谢的关键酶。虽然已克隆并在重组大肠杆菌中表达，但重组蛋白是在包涵体中以失活形式合成的。然而，我们已经开发了一种新的方法，通过在酸性条件下重折叠获得活性形式的重组af - th。将酶纯化至均匀并结晶。x射线晶体分析表明，该晶体衍射分辨率为2.15 A，属于p31或p32空间群，晶胞参数A = b= 92.1, c = 232.6 A。需要进一步的分析，如多/单异常色散方法来确定af - th结构。

This study has been performed to progress our knowledge about the enzymes involved in the dissimilatory inorganic sulfur metabolism in the sulfur-oxidizing bacteria. We have isolated and identified the gene encodingtetrathionate hydrolase (Af-tth) which is the key enzyme in the metabolism in Acidithiobacillus ferrooxidans. Although Af-tthwas cloned and expressed in recombinant Escherichia coli, the recombinant proteins were synthesized in inclusion bodies as an inactive form. However, we have developed a new method to obtain the recombinant Af-Tth in an active form by refolding under acidic conditions. The enzyme was purified to homogeneity and crystallized. X-ray crystallographic analysis showed that the crystal diffracted to 2.15 A resolution and belongs to space group P31or P32with unit-cell parameters a = b= 92.1, c = 232.6 A. Further analyses such as multiple/single anomalous dispersion methods are required to determine the Af-Tth structure.