Production, crystallization and preliminary crystallographic analysis ofAllochromatium vinosumthiosulfate dehydrogenase TsdA, an unusual acidophilicc-type cytochrome

Abstract

The ability to perform the very simple oxidation of two molecules of thiosulfate to tetrathionate is widespread among prokaryotes. Despite the prevalent occurrence of tetrathionate formation and its well documented significance within the sulfur cycle, little is known about the enzymes that catalyze the oxidative condensation of two thiosulfate anions. To fill this gap, the thiosulfate dehydrogenase (TsdA) enzyme from the purple sulfur bacteriumAllochromatium vinosumwas recombinantly expressed inEscherichia coli, purified and crystallized, and a crystallographic data set was collected. The crystals belonged to the monoclinic space groupC2, with unit-cell parametersa= 79.2,b= 69.9,c= 57.9 Å, β = 129.3°, contained one monomer per asymmetric unit and diffracted to a resolution of 1.98 Å.

A novel bacterial thiosulfate oxidation pathway provides a new clue about the formation of zero-valent sulfur in deep sea

“…The apparent molecular mass of purified E. flavus 21–3 TsdA is 36 KD (Supplementary Fig. S3b ), larger than the homologous protein in A. vinosum (27 KD) [ 60 ]. The UV–Visible electronic absorption spectrum of the purified recombinant protein has an obvious absorption at 410 nm (Supplementary Fig.…”

X-ray–induced photoreduction of heme metal centers rapidly induces active-site perturbations in a protein-independent manner

“…For heme proteins, there are essentially three reported cases: myoglobin (10,(19)(20)(21), cytochrome c peroxidase, and horseradish peroxidase (8,9). For a more general approach to study reduction kinetics, a set of six diverse heme proteins was selected; metmyoglobin from horse heart (hhMb), a diheme c protein, thiosulfate dehydrogenase from Allochromatium vinosum (AvTsdA) (22)(23)(24)(25), a B-class dye-decolorizing peroxidase (KpDyP) (26), chlorite dismutases from Nitrospira defluvii (NdCld) (27) and from Cyanothece sp. PCC7425 (CCld) (28), and a coproheme decarboxylase from Listeria monocytogenes (LmChdC) (29).…”

Thiosulfate Dehydrogenase (TsdA) from Allochromatium vinosum

“…Crystallization and Data Collection-TsdA crystallization has been reported previously (32). In summary, TsdA at a concentration of 8 mg ml Ϫ1 in 20 mM BisTris-HCl, pH 6.5, crystallized in a condition comprising 23.5% (w/v) PEG 3350, 0.2 M (NH 4 ) 2 SO 4 , 0.1 M BisTris, pH 6.28, and 0.1 M NaI (as additive).…”

“…At A 600 of 0.6, the culture was switched to 25°C, and the cells were harvested after an additional 16 -20 h. Cells were resuspended in 50 mM BisTris-HCl buffer, pH 6.5, and lysed by sonication. After the removal of insoluble cell material by centrifugation (10,000 ϫ g for 25 min at 4°C), TsdA wild type or TsdA mutant proteins were purified by Strep-Tactin affinity chromatography and gel filtration as described before (32).…”