Abstract
Sulfur oxygenase reductases (SORs) are present in thermophilic and mesophilic archaea and bacteria, and catalyze oxygen-dependent oxygenation and disproportionation of elemental sulfur. SOR has a hollow, spherical homo-24-mer structure and reactions take place at active sites inside the chamber. The crystal structures of SORs from Acidianus species have been reported. However, the states of the active site components (mononuclear iron and cysteines) and the entry and exit paths of the substrate and products are still in dispute. Here, we report the biochemical and structural characterizations of SORs from the thermoacidophilic archaeon Sulfurisphaera tokodaii (StSOR) and present high-resolution structures determined by X-ray crystallography and cryogenic electron microscopy (cryo-EM). The crystal structure of StSOR was determined at 1.73 Å resolution. At the catalytic center, iron is ligated to His86, His90, Glu114, and two water molecules. Three conserved cysteines in the cavity are located 9.5–13 Å from the iron and were observed as free thiol forms. A mutational analysis indicated that the iron and one of the cysteines (Cys31) were essential for both activities. The cryo-EM structure was determined at 2.24 Å resolution using an instrument operating at 200 kV. The two structures determined by different methodologies showed similar main chain traces, but the maps exhibited different features at catalytically important components. A possible role of StSOR in the sulfur metabolism of S. tokodaii (an obligate aerobe) is discussed based on this study. Given the high resolution achieved in this study, StSOR was shown to be a good benchmark sample for cryo-EM.
Highlights
In addition to geochemical activities, archaea and bacteria play significant roles in both oxidation and reduction of sulfur compounds in the global sulfur cycle
On sodium dodecyl sulfate (SDS)-denatured polyacrylamide gel electrophoresis (PAGE), a major single band appeared at a position of apparent molecular mass of 38 kDa, which was comparable to the predicted molecular mass of the recombinant protein (35.7 kDa)
4.1 Substrate entry and exit pathways The pathways for substrate entry and exit have been major topics of discussion regarding sulfur oxygenase reductase (SOR) structures because catalysis takes place inside the hollow chamber that is separated from the cytoplasmic environment (Li et al, 2008; Veith et al, 2011)
Summary
In addition to geochemical activities, archaea and bacteria play significant roles in both oxidation and reduction of sulfur compounds in the global sulfur cycle. Oxidation of inorganic sulfur compounds under anaerobic conditions is the most important energyyielding reaction for microorganisms living in those critical environments (Ghosh and Dam, 2009; Offre et al, 2013). Solfataras, volcanic hot springs and volcanic environments all over the world, thermophilic archaea and bacteria grow under these high temperature and acidic conditions (Schönheit and Schäfer, 1995). Such thermoacidophiles belonging to the order Sulfolobales show biotechnological possibility for application to biomining, where metal sulfides in insoluble ore are recovered through the sulfur-metabolizing process of the organisms (Mangold et al, 2011; Urbieta et al, 2017)
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