Atrazine is the most widely used herbicide by corn and sorghum growers. Atrazine
chlorohydrolase (AtzA) dechlorinates atrazine, producing non-toxic and non-phytotoxic
hydroxyatrazine. In this study, we report the cloning and initial experiments for partial
characterization of mutant AtzA (Colin Scott, Colin J. Jackson, Chris W. Coppin, et al.
2009. Catalytic Improvement and Evolution of Atrazine Chlorohydrolase. Appl. Environ.
Microbiol. 75(7):2184-2191). A plasmid vector was designed for ease of purification of
the enzyme with the help of a his-tag and to ensure high expression of soluble protein in
recombinant E.coli. The specific activity and substrate specificity of the purified mutant
AtzA was then compared to the wild type enzyme. It was observed from these initial
studies that the mutant enzyme had somewhat similar characteristics to the wild type
Further, the current study also describes immobilization recombinant E. coli cells
expressing the wild type atrazine chlorohydrolase in a silica/polymer porous gel. This
novel recombinant enzyme-based method utilizes both adsorption and degradation to
remove atrazine from water. A combination of silica nanoparticles (Ludox TM40),
alkoxides, and an organic polymer were used to synthesize a porous gel. Gel curing
temperatures of 23°C or 45°C were used to either maintain cell viability or to render the
cells non-viable, respectively. The enzymatic activity of the encapsulated viable and nonviable
cells was high and extremely stable over the time period analyzed. At room
temperature, the encapsulated non-viable cells maintained a specific activity between (0.44 ± 0.06) μmol/g-min and (0.66 ± 0.12) μmol/g-min for up to 4 months, comparing
well with free, viable cells specific activities (0.61 ± 0.04 μmol/g-min). Gels cured at 45°C
had excellent structural rigidity and contained few viable cells, making these gels
potentially compatible with water treatment facility applications.
University of Minnesota M.S. thesis. January 2012. Major:Biochemistry, Molecular Biology and Biophysics. Advisors: Lawrence P. Wackett & Ping Wang. 1 computer file (PDF); viii, 85 pages.
Biodegradation of atrazine by atrazine chlorohydrolase: characterization of mutant enzyme and immobilization system for water purification..
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