tailieunhanh - Báo cáo Y học: Cloning, expression and characterization of a gene encoding nitroalkane-oxidizing enzyme from Streptomyces ansochromogenes

A nitroalkane-oxidizing enzyme gene (naoA) was cloned from a genomic DNA library ofStreptomyces ansochromo-genes7100. The deduced protein (NaoA) of this gene con-tains 363 amino acids and has high similarity to several nitroalkane-oxidizing enzymes from various subcloned intoanexpressionvector pET23b and overexpressed inEscherichia coliBL21(DE3). The protein was then purified, and its characteristics were studied. Experimental results showed that NaoA can con-vert 1-nitropropane, 2-nitropropane and nitroethane into the corresponding carbonyl compounds | Eur. J. Biochem. 269 6302-6307 2002 FEBS 2002 doi Cloning expression and characterization of a gene encoding nitroalkane-oxidizing enzyme from Streptomyces ansochromogenes Jihui Zhang Wenbo Ma and Huarong Tan Institute of Microbiology Chinese Academy of Sciences Beijing China A nitroalkane-oxidizing enzyme gene naoA was cloned from a genomic DNA library of Streptomyces ansochromo-genes 7100. The deduced protein NaoA of this gene contains 363 amino acids and has high similarity to several nitroalkane-oxidizing enzymes from various micro-organisms. The naoA gene was subcloned into an expression vector pET23b and overexpressed in Escherichia coli BL21 DE3 . The protein was then purified and its characteristics were studied. Experimental results showed that NaoA can convert 1-nitropropane 2-nitropropane and nitroethane into the corresponding carbonyl compounds. The optimal pH and temperature for NaoA was found to be pH 7-8 and 48-56 C respectively. The Km of NaoA for nitroethane is w mM. NADH and nitro blue tetrazolium are strong inhibitors of NaoA and thiol compounds and superoxide dismutase partially inhibit the enzyme activity. Therefore superoxide may be an essential intermediate in the oxidation of nitroalkane by NaoA. Keywords enzymatic properties expression gene cloning nitroalkane-oxidizing enzyme Streptomyces. Nitroalkane compounds are widely used in chemical industry as intermediates solvents and fuel for rockets 1 and are released in large quantities into the environment. Meanwhile certain micro-organisms and many leguminous plants produce nitroalkane compounds 2 . These materials are hazardous and can result in environmental contamination. So the conversion of nitro groups by biocatalysts is useful in industry as well as in environmental conservation. Enzymes that can convert nitroalkanes into less harmful species have been purified and characterized from micro-organisms. They include 2-nitropropane dioxygenase from .

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