Reduction of Hexavalent Chromium by Cell-Free Extract of Bacillus sphaericus AND 303 Isolated from Serpentine Soil

Abstract

Cell-free extracts (CFEs) of chromium-resistant bacterium Bacillus sphaericus AND 303 isolated from serpentine soil of Andaman, India reduced Cr(VI) in in vitro condition, and the reductase activity was solely localized in the soluble cell-fractions (S12, S32, and S150). The enzyme was constitutive as the CFEs from cells grown in Cr(VI)-free and Cr(VI)-containing media reduced a more or less equal amount of Cr(VI). Optimum Cr(VI) reductase activity was obtained at an enzyme (S150) concentration equivalent to 4.56 mg protein/mL, 300 microM: Cr(VI) and pH 6.0 after 30 min incubation at 30 degrees C. The enzyme was heat labile; 80% of its activity was lost when exposed at 70 degrees C for 15 min. Kinetics of Cr(VI) reductase activity fit well with the linearized Lineweaver-Burk plot and showed a V(max) of 1.432 micromol Cr(VI)/mg protein/min and K(m) of 158.12 microM: Cr(VI). The presence of additional electron donors accelerated Cr(VI) reductase activity of CFE, and an increase of 28% activity over control was recorded with 1.0 microM: NADH. Heavy metal ions such as Ni(II), Cu(II), and Cd(II) were strong inhibitors of Cr(VI) reductase unlike that of 100 microM: Co(II), which retained 93% activity over control.

Treatment of Chromium Pollution by the Reductase Enzyme Generated by a Chromium-Resistant Bacterium Isolated in Dewatering Sludge

“…The observation was made with the pH, having the maximum activity at pH 7. Similar observations of the influence of temperature and pH in the enzyme activity have been made in the literature (Pal et al, 2005;Desai et al, 2008). Suziki et al (1992) reported that some bacteria were known to use chromate as a terminal electron acceptor employing membrane bound enzymes, while others use soluble enzymes.…”

“…and Microbacterium sp. were recently investigated (Pal et al, 2005;Thacker et al, 2006;Soni et al, 2013). The enzyme activity reported by Desai and his coworkers (2008) were lower compared to the values obtained in our experiments at the same Cr (VI) concentration, suggesting that our strain is a higher reductase producing bacterium among Bacillus strains reported up to date (Table 2).…”

“…In our experiments, higher Cr (VI) concentration (3.4 mM) caused decrease in growth rate when compared to growth at lower concentrations; this was ascribed to the toxic effect of Cr (VI). Toxic effect of Cr (VI) on the cell growth at higher concentration has been mentioned in the literature (Pal et al, 2005;Cheng and Li, 2009). Further, the total chromium remained almost the same at different Cr (VI) concentrations and this suggests that the decrease of Cr (VI) concentration in the supernatant was the effect of reduction of Cr (VI) to Cr (III) and not adsorption (Pal et al, 2005).…”

“…The enzyme assays were performed as previously described Pal et al (2005). The reaction mixture containing Cr (VI) at the final concentration of 0.1 mM and 2 mM, glucose (0.2 µM) in 50 mM phosphate buffer and 1 ml of enzyme reductase.…”

Hexavalent chromium reduction by immobilized cells of Bacillus sphaericus AND 303

“…Growing bacterial cells as well as their cell-free extracts reduced Cr(VI) under aerobic or anaerobic condition or both. While aerobic chromate reduction by bacteria is associated with soluble cellular proteins utilizing endogenous electron reserves or NADH as the electron donor (Park et al 2000;Pal et al 2005), anaerobic reduction is membrane bound where Cr(VI) serves as the terminal electron acceptor through respiratory chains involving cytochromes Shen and Wang 1994). Bacterial cells and enzymes immobilized in different polymer matrices such as agar-agar, agarose, polyacrylamide, calcium alginate, diatomite, polyvinyl alcohol, etc.…”