Structure and function prediction of arsenate reductase from Deinococcus indicus DR1.

Arsenic prevalence in the environment impelled many organisms to develop resistance over the course of evolution. Tolerance to arsenic, either as the pentavalent [As(V)] form or the trivalent form [As(III)], by bacteria has been well studied in prokaryotes, and the mechanism of action is well defined. However, in the rod-shaped arsenic tolerant Deinococcus indicus DR1, the key enzyme, arsenate reductase (ArsC) has not been well studied. ArsC of D. indicus belongs to the Grx-linked prokaryotic arsenate reductase family. While it shares homology with the well-studied ArsC of Escherichia coli having a catalytic cysteine (Cys 12) and arginine triad (Arg 60, 94, and 107), the active site of D.indicus ArsC contains four residues Glu 9, Asp 53, Arg 86, and Glu 100, and with complete absence of structurally equivalent residue for crucial Cys 12. Here, we report that the mechanism of action of ArsC of D. indicus is different as a result of convergent evolution and most likely able to detoxify As(V) using a mix of positively- and negatively-charged residues in its active site, unlike the residues of E. coli. This suggests toward the possibility of an alternative mechanism of As (V) degradation in bacteria.

Pentavalent arsenate reductase activity in cytosolic fractions of Pseudomonas sp., isolated from arsenic-contaminated sites of Tezpur, Assam.

Pentavalent arsenate reductase activity was localized and characterized in vitro in the cytosolic fraction of a newly isolated bacterial strain from arsenic-contaminated sites. The bacterium was gram negative, rod-shaped, nonmotile, non-spore-forming, and noncapsulated, and the strain was identified as Pseudomonas sp. DRBS1 following biochemical and molecular approaches. The strain Pseudomonas sp. DRBS1 exhibited enzymatic machinery for reduction of arsenate(V) to arsenite(III). The suspended culture of the bacterium reduced more than 97% of As(V) (40-100 mM) to As(III) in 48 h. The growth rate and total cellular yield decreased in the presence of higher concentration of arsenate. The suspended culture repeatedly reduced 10 mM As(V) within 5 h up to five consecutive inputs. The cell-free extracts reduced 86% of 100 microM As(V) in 40 min. The specific activity of arsenate reductase enzyme in the presence of 100 microM arsenate is 6.68 micromol/min per milligram protein. The arsenate reductase activity is maximum at 30 degrees C and at pH 5.2. The arsenate reductase activity increased in the presence of electron donors like citrate, glucose, and galactose and metal ions like Cd(+2), Cu(+2), Ca(+2), and Fe(+2). Selenate as an electron donor also supports the growth of strain DRBS1 and significantly increased the arsenate reduction.

Respiratory arsenate reductase as a bidirectional enzyme.

The haloalkaliphilic bacterium Alkalilimnicola ehrlichii is capable of anaerobic chemolithoautotrophic growth by coupling the oxidation of arsenite (As(III)) to the reduction of nitrate and carbon dioxide. Analysis of its complete genome indicates that it lacks a conventional arsenite oxidase (Aox), but instead possesses two operons that each encode a putative respiratory arsenate reductase (Arr). Here we show that one homolog is expressed under chemolithoautotrophic conditions and exhibits both arsenite oxidase and arsenate reductase activity. We also demonstrate that Arr from two arsenate respiring bacteria, Alkaliphilus oremlandii and Shewanella sp. strain ANA-3, is also biochemically reversible. Thus Arr can function as a reductase or oxidase. Its physiological role in a specific organism, however, may depend on the electron potentials of the molybdenum center and [Fe-S] clusters, additional subunits, or constitution of the electron transfer chain. This versatility further underscores the ubiquity and antiquity of microbial arsenic metabolism.