Hexavalent chromate reductase activity in cytosolic fractions of Pseudomonas sp. G1DM21 isolated from Cr(VI) contaminated industrial landfill

Hexavalent chromate reductase activity was localized and characterized in vitro in cytosolic fraction of a newly isolated Pseudomonas sp. G1DM21. The suspended culture of the bacterium reduced 99.7% of 500 μM Cr(VI) and 93.06% of 1000 μM Cr(VI) in 48 h. The suspended culture repeatedly reduced 100 μM Cr(VI) within 6 h up to four consecutive inputs. The permeabilized cells of the bacterium reduced 92% of 100 μM Cr(VI) within 6 h. The cell-free extracts (CFE) reduced 90% of 100 μM Cr(VI) in 120 min. The Km and Vmax determined for chromate reductase activity in the CFE were 175 μM Cr(VI) and 1.6 μmoles/min/mg of protein, respectively, the Km and Vmax determined in the presence of 0.5 mM NADH were 150 μM Cr(VI) and 2.0 μmoles/min/mg of protein, respectively. Hexavalent chromate reductase activity was maximum at 30 °C and pH 7.0. Relative molecular mass (Mr) of the native Cr(VI) reductase in the cytosolic fraction was estimated as 61.7 kDa. The Cr(VI) reductase activity increased in the presence of metal ions like Cu2+, Mg2+, Na+ and electron donors like citrate, succinate, acetate and was significantly inhibited in the presence of metal ions like Hg2+, Ag+, Cd2+, disulfide reducers like 2-mercaptoethanol, while the respiratory inhibitors had a minute effect on the activity. Scanning probe atomic force microscopy (AFM) analysis indicates that exposure of Pseudomonas sp. G1DM21 to 1 mM Cr(VI) for 24 h, leads to an increase in cell length and height.