Chromium (VI) (chromate) is a highly soluble, toxic, and mutagenic contaminant resulting from numerous industrial processes, and is present in soil and groundwater worldwide. Efforts toward bioremediation of chromate to Cr (III) using enzymatic action by select bacteria have been pursued in order to diminish toxicity and solubility. Previously reported chromate reductases were found in Pseudomonas species which are known for their toxicity resistance to chromium, silver, mercury, hydrocarbons, and other pollutants. (1,2) Using a modified Environmental Protection Agency (EPA) colorimetric analysis, a novel bacteria has been identified that has chromate reductase activity. Partial purification has been achieved through centrifugation and anion exchange chromatography (DEAE). The DEAE resulted in two possible enzymes with reductase activity. The one with higher activity was retained for further study. The partial purification has been yielded some characterization of the enzyme. The enzyme is under 50 kDa, dependent upon NADH, has an optimal temperature of 40°C, and denatures at 70°C. Elution from the DEAE column appeared early in the salt gradient, suggesting a slightly negative overall charge. Isolation continues by high performance liquid chromatography with reverse phase hydrophobic column. Obtaining a purified enzyme will allow full kinetic studies and possible sequencing for comparison to reductase homologs. References: 1. Park, C., M. Keyhan, B. Wielinga, S. Fendorf, and A. Martin. 2000. Purification to homogeneity and characterization of a novel Pseudomonas putida chromate reductase. Appl. Environ. Microbiol. 66:1788-1795 2. Suzuki, T., Miyata, H. Horitsu, K. Kawai, K. Takamizawa, Y. Tai, and M. Okazaki. 1992. NAD(P)H-dependent Chromium (VI) reductase of Pseudomonas ambigua G-1: a Cr(v) intermediate is formed during the reduction of Cr(VI) to Cr(III). J. Bacteriol. 174:5340-5345.

Isolation of a chromate reductase from Pseudomonas veronii

251st National Meeting (2016)

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